From Jane Doe to Sofia: DNA Extraction Protocol from Bones and Teeth without Liquid Nitrogen for Identifying Skeletal Remains.

DNA analysis plays a crucial role in forensic investigations, helping in criminal cases, missing persons inquiries, and archaeological research. This study focuses on the DNA concentration in different skeletal elements to improve human identification efforts. Ten cases of unidentified skeletal remains brought to the Institute of Forensic Medicine in Timisoara, Romania, underwent DNA analysis between 2019 and 2023. The results showed that teeth are the best source for DNA extraction as they contain the highest concentration of genetic material, at 3.68 ng/µL, compared to the petrous temporal bone (0.936 ng/µL) and femur bone (0.633 ng/µL). These findings highlight the significance of teeth in forensic contexts due to their abundant genetic material. Combining anthropological examination with DNA analysis enhances the understanding and precision of identifying human skeletal remains, thus advancing forensic science. Selecting specific skeletal elements, such as the cochlea or teeth, emerges as crucial for reliable genetic analyses, emphasizing the importance of careful consideration in forensic identification procedures. Our study concludes that automated DNA extraction protocols without liquid nitrogen represent a significant advancement in DNA extraction technology, providing a faster, more efficient, and less labor-intensive method for extracting high-quality DNA from damaged bone and tooth samples.

A Retrospective Assessment of Sputum Samples and Antimicrobial Resistance in COVID-19 Patients.

Data on bacterial or fungal pathogens and their impact on the mortality rates of Western Romanian COVID-19 patients are scarce. As a result, the purpose of this research was to determine the prevalence of bacterial and fungal co- and superinfections in Western Romanian adults with COVID-19, hospitalized in in-ward settings during the second half of the pandemic, and its distribution according to sociodemographic and clinical conditions. The unicentric retrospective observational study was conducted on 407 eligible patients. Expectorate sputum was selected as the sampling technique followed by routine microbiological investigations. A total of 31.5% of samples tested positive for Pseudomonas aeruginosa, followed by 26.2% having co-infections with Klebsiella pneumoniae among patients admitted with COVID-19. The third most common Pathogenic bacteria identified in the sputum samples was Escherichia coli, followed by Acinetobacter baumannii in 9.3% of samples. Commensal human pathogens caused respiratory infections in 67 patients, the most prevalent being Streptococcus penumoniae, followed by methicillin-sensitive and methicillin-resistant Staphylococcus aureus. A total of 53.4% of sputum samples tested positive for Candida spp., followed by 41.1% of samples with Aspergillus spp. growth. The three groups with positive microbial growth on sputum cultures had an equally proportional distribution of patients admitted to the ICU, with an average of 30%, compared with only 17.3% among hospitalized COVID-19 patients with negative sputum cultures (p = 0.003). More than 80% of all positive samples showed multidrug resistance. The high prevalence of bacterial and fungal co-infections and superinfections in COVID-19 patients mandates for strict and effective antimicrobial stewardship and infection control policies.

Connections between Diabetes Mellitus and Metabolic Syndrome and the Outcome of Cardiac Dysfunctions Diagnosed during the Recovery from COVID-19 in Patients without a Previous History of Cardiovascular Diseases.

(1) Background: Throughout the COVID-19 pandemic, it became obvious that individuals suffering with obesity, diabetes mellitus (T2DM), and metabolic syndrome (MS) frequently developed persisting cardiovascular complications, which were partially able to explain the onset of the long-COVID-19 syndrome. (2) Methods: Our aim was to document, by transthoracic echocardiography (TTE), the presence of cardiac alterations in 112 patients suffering from post-acute COVID-19 syndrome and T2DM, MS, and/or obesity, in comparison to 91 individuals without metabolic dysfunctions (MD); (3) Results: in patients with MD, TTE borderline/abnormal left (LVF) and/or right ventricular function (RVF), alongside diastolic dysfunction (DD), were more frequently evidenced, when compared to controls (p ˂ 0.001). Statistically significant associations between TTE parameters and the number of factors defining MS, the triglyceride-glucose (TyG) index, the severity of the SARS-CoV-2 infection, and the number of persisting symptoms (p ˂ 0.001) were noted. Significant predictive values for the initial C-reactive protein and TyG index levels, both for the initial and the 6-month follow-up levels of these TTE abnormalities (p ˂ 0.001), were highlighted by means of a multivariate regression analysis. (4) Conclusions: in diabetic patients with MS and/or obesity with comorbid post-acute COVID-19 syndrome, a comprehensive TTE delineates various cardiovascular alterations, when compared with controls. After 6 months, LVF and RVF appeared to normalize, however, the DD-although somewhat improved-did persist in approximately a quarter of patients with MD, possibly due to chronic myocardial changes.

Molecular Monitoring of Allogeneic Stem Cell Transplantation in Fanconi Anemia.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (HSCT) is the treatment of choice in patients with Fanconi anemia (FA). The aim of our study is to evaluate the impact and benefits of allogenic matched donor HSCT in a case of a 12 year-old girl with FA, who displayed good clinical evolution following 2 months post-transplantation. METHODS: In the pre-transplant phase, reference blood samples from the donor and recipient were collected on EDTA. The DNA from blood samples was extracted using an automated Maxwell® 48 RSC instrument (Promega, USA) with the Maxwell® RSC Whole blood DNA kit (Promega, USA). For DNA quantification, the PowerQuant System kit (Promega, USA) was used with the ABI 7500 Real-time PCR system (Applied Biosystems, USA). The amplification of the short tandem repeat markers was performed using the 24plex Investigator QS kit (Qiagen, Germany) on a ProFlex PCR System. Furthermore, the PCR products were separated and detected on an ABI 3500 Genetic Analyzer (Applied Biosytems, USA). RESULTS: Thirty days post transplantation, a complete chimerism (CC) was achieved with a full replacement by do-nor derived hematopoietic cells. Sixty days post transplantation, the CC status was maintained with improvement of hematological findings. CONCLUSIONS: In FA, chimerism monitoring after HSCT provides useful information regarding engraftment or possibility of post-transplantation complications such as graft versus host disease.

The Impact of Metabolic Syndrome and Obesity on the Evolution of Diastolic Dysfunction in Apparently Healthy Patients Suffering from Post-COVID-19 Syndrome.

(1) Background: Coronavirus disease 2019 (COVID-19) has a worse prognosis in individuals with obesity and metabolic syndrome (MS), who often develop cardiovascular complications that last throughout recovery. (2) Methods: This study aimed to analyze the evolution of diastolic dysfunction (DD), assessed by transthoracic echocardiography (TTE), in 203 individuals with and without obesity and/or MS diagnosed with post-COVID-19 syndrome. (3) Results: DD was frequently diagnosed in patients with MS and obesity, but also in those without obesity (62.71% and 56.6%, respectively), in comparison to 21.97% of subjects without MS (p ˂ 0.001). Almost half of the patients with obesity and MS had more severe DD (types 2 and 3). As for evolution, the prevalence and severity of DD, particularly types 1 and 2, decreased gradually, in parallel with the improvement of symptoms, progress being more evident in subjects without MS. DD of type 3 did not show a significant reduction (p = 0.47), suggesting irreversible myocardial damages. Multivariate regression analysis indicated that the number of MS factors, the severity of initial pulmonary injury, and protein C levels could explain DD evolution. (4) Conclusions: DD was commonly diagnosed in individuals with post-COVID-19 syndrome, particularly in those with MS and obesity. After 6 months, DD evolution, excepting that of type 3, showed a significant improvement, mostly in patients without MS.

Postmortem MicroRNA Signatures as Predictive Markers in SARS-CoV-2 Infection.

BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disease that emerged in December 2019 from Wuhan, China, has led to a worldwide outbreak that has resulted in 234,809,103 confirmed cases and caused more than 4,800,375 deaths worldwide. MicroRNAs could be involved in the SARS-CoV-2 infection, but not many studies have been performed to explore this in postmortem cases. The purpose of our study is to evaluate the postmortem expression of microRNA-6501-5p, microRNA-5695, and microRNA-29b-3p from bronchial secretions in positive and negative SARS-CoV-2 deaths and to evaluate their usefulness as predictive biomarkers in the evolution of SARS-CoV-2 infection. METHODS: During the autopsy procedure on 61 "suspected" deaths at the Institute of Forensic Medicine in Timisoara, Romania, bronchial secretions were collected to detect SARS-CoV-2 infection postmortem. After the RT-PCR analysis, 44 SARS-CoV-2 cases were detected positive, while 17 cases were SARS-CoV-2 negative, which were considered as controls. RESULTS: From the panel of microRNAs, microRNA-6501-5p, microRNA-5695, and microRNA-29b-3p were upregulated in SARS-CoV-2 cases and down-regulated in non-SARS-CoV-2 cases. CONCLUSIONS: We concluded that using a panel of microRNAs as biomarkers in SARS-CoV-2 infection could aid in an early evaluation of the evolution of SARS-CoV-2-infected patients.

SARS-CoV-2: An Overview of the Genetic Profile and Vaccine Effectiveness of the Five Variants of Concern.

With the onset of the COVID-19 pandemic, enormous efforts have been made to understand the genus SARS-CoV-2. Due to the high rate of global transmission, mutations in the viral genome were inevitable. A full understanding of the viral genome and its possible changes represents one of the crucial aspects of pandemic management. Structural protein S plays an important role in the pathogenicity of SARS-CoV-2, mutations occurring at this level leading to viral forms with increased affinity for ACE2 receptors, higher transmissibility and infectivity, resistance to neutralizing antibodies and immune escape, increasing the risk of infection and disease severity. Thus, five variants of concern are currently being discussed, Alpha, Beta, Gamma, Delta and Omicron. In the present review, a comprehensive summary of the following critical aspects regarding SARS-CoV-2 has been made: (i) the genomic characteristics of SARS-CoV-2; (ii) the pathological mechanism of transmission, penetration into the cell and action on specific receptors; (iii) mutations in the SARS-CoV-2 genome; and (iv) possible implications of mutations in diagnosis, treatment, and vaccination.

Autopsy Findings and Inflammatory Markers in SARS-CoV-2: A Single-Center Experience.

Purpose: The systemic inflammatory response related to COVID-19 can be easily investigated in living patients. Unfortunately, not every biomarker is suitable for postmortem analysis since several factors may interfere. The aim of this study was to summarize key histopathological findings within each organ system due to COVID-19 and to assess if serological inexpensive and widely available biomarkers such as CRP, IL-6, fibrinogen and d-Dimers, associated with adverse outcomes in COVID-19, can be implemented in a post-mortem assessment. Patients and Methods: A total of 60 subjects divided in 2 groups were included. All subjects died outside a hospital setting and therefore did not receive specific or symptomatic therapies that could have modulated the inflammatory response. The first group included 45 subjects in which mandatory autopsy was performed in order to establish the cause of death and macroscopic examination of the lungs was highly suggestive of SARS-CoV-2 infection. As controls (Group 2), 20 subjects who died from polytrauma in high velocity car accidents and suicide were selected. Bronchial fluids collected during the autopsy procedure were used for the RT-PCR diagnosis of SARS-CoV-2 and serum samples were sent for analysis of IL-6, CRP, d-Dimers and fibrinogen. Results: Compared with the control group, the subjects of the COVID-19 group were older (59±19.5 vs.38±19.15 years, p=0.0002) and had more underlying comorbidities such as hypertension (60% vs 35%, p=0.06) or were overweight (53.3% vs 30%, p=0.08). The levels of CRP, IL-6, fibrinogen and d-Dimers in postmortem plasma samples were significantly higher in COVID-19 subjects than in control group (p< 0.0001). Moreover, the level of IL-6 was significantly higher in overweight patients (r=0.52, P<0.001). In all COVID-19 subjects, the histological examination revealed features corresponding to the exudative and/or proliferative phases of diffuse alveolar damage. Large pulmonary emboli were observed in 7 cases. Gross cardiac enlargement with left ventricular hypertrophy was observed in 19 cases. The most frequent pathological finding of the central nervous system was acute/early-subacute infarction. Conclusion: Due to the complexity of the inflammatory response, we postulate that a combination of biomarkers, rather than a single laboratory parameter, might be more effective in obtaining a reliable postmortem COVID-19 diagnosis.

Oleogel Formulations for the Topical Delivery of Betulin and Lupeol in Skin Injuries-Preparation, Physicochemical Characterization, and Pharmaco-Toxicological Evaluation.

The skin integrity is essential due to its pivotal role as a biological barrier against external noxious factors. Pentacyclic triterpenes stand as valuable plant-derived natural compounds in the treatment of skin injuries due to their anti-inflammatory, antioxidant, antimicrobial, and healing properties. Consequently, the primary aim of the current investigation was the development as well as the physicochemical and pharmaco-toxicological characterization of betulin- and lupeol-based oleogels (Bet OG and Lup OG) for topical application in skin injuries. The results revealed suitable pH as well as organoleptic, rheological, and textural properties. The penetration and permeation of Bet and Lup oleogels through porcine ear skin as well as the retention of both oleogels in the skin were demonstrated through ex vivo studies. In vitro, Bet OG and Lup OG showed good biocompatibility on HaCaT human immortalized cells. Moreover, Bet OG exerted a potent wound-healing property by stimulating the migration of the HaCaT cells. The in ovo results demonstrated the non-irritative potential of the developed formulations. Additionally, the undertaken in vivo investigation indicated a positive effect of oleogels treatment on skin parameters by increasing skin hydration and decreasing erythema. In conclusion, oleogel formulations are ideal for the local delivery of betulin and lupeol in skin disorders.

Alterations of Left Ventricular Function Persisting during Post-Acute COVID-19 in Subjects without Previously Diagnosed Cardiovascular Pathology.

(1) Background: Coronavirus infection (Covid-19) has emerged as a severe medical condition, associated with high pulmonary morbidity and often with cardiovascular (CV) complications. This study aims to evidence the persistence of left ventricular (LV) systolic function (LV-SF) alterations and diastolic dysfunction (DD) in COVID-19 patients without history of cardiovascular (CV) diseases by transthoracic echocardiography (TTE). (2) Methods: 125 patients, aged under 55 years, hospitalized during the first outbreak of Covid-19 for moderate pneumonia, underwent a comprehensive cardiologic examination and TTE at 6-10 weeks after discharge. Their initial in-hospital laboratory data and thorax computer tomography (TCT) were accessed from the electronic database of the hospital. (3) Results: with TTE, we documented alterations of LV-SF and DD in 8.8% of patients and in 16.8% only patterns of DD, statistically correlated with the initial levels of creatin-kinase (CK-MB) and inflammatory factors. Multivariate regression analysis evidenced that CK-MB levels, age, and body mass index (BMI) are responsible for 65% of LV-SF decrease. (4) Conclusions: Alterations of LV-SF and DD are frequent in post-acute COVID-19 infection and are responsible for the persistence of symptoms. Elevated myocardial necrosis markers during the acute phase seem to predict subsequent alteration of cardiac performance.

Evidence of Pulmonary Hypertension after SARS-CoV-2 Infection in Subjects without Previous Significant Cardiovascular Pathology.

BACKGROUND: Coronavirus (Covid-19) infection represents a serious medical condition, often associated with cardiovascular complications, pulmonary hypertension (PH), and right ventricle dysfunction (RVD). The aim of this study is to show, by means of transthoracic echocardiography (TTE), the presence of an increased estimated systolic pressure in the pulmonary artery (esPAP) and altered right ventricular global longitudinal strain (RV-GLS) in patients without history of PH. METHODS: In a group of 91 patients, aged under 55 years, hospitalized for a moderate Covid-19 infection, a thorough cardiologic and TTE examination were performed two months after discharge. Their initial thorax computer-tomography (TCT) images and laboratory data were accessed from the electronic data base of the hospital. RESULTS: We observed an increased prevalence of PH (7.69%) and RVD (10.28%), significantly correlated with the initial levels of the TCT score and inflammatory factors (p ˂ 0.001), but borderline changes were observed in more patients. Multivariate regression analysis showed that these factors and RV-GLS explain 89.5% of elevated esPAP. CONCLUSIONS: In COVID-19 infection, PH and RVD are common complications, being encountered after the recovery even in moderate cases. It appears to be a connection between their severity and the extent of the initial pulmonary injury and of the inflammatory response.

Molecular Testing of SARS-CoV-2 Infection from Blood Samples in Disseminated Intravascular Coagulation (DIC) and Elevated D-Dimer Levels.

BACKGROUND: In 2020, the SARS-CoV-2 virus spread worldwide and infected more that 10 million people, causing more than 500,000 deaths worldwide. The infection has systemic effects on the respiratory and cardiovascular systems; thus, patients can present a variety of symptoms from asymptomatic to rapid deaths. In this paper, we present the first case of post-mortem SARS-CoV-2 molecular testing in Western part of Romania in a deceased with disseminated intravascular coagulation (DIC) and elevated D-dimer levels. METHODS: During the autopsy which took place at the Institute of Forensic Medicine from Timisoara, Romania, blood sample was collected in a vacutainer with EDTA and sent to the Laboratory of Forensic Genetics from Victor Babes University of Medicine and Pharmacy, Timisoara, Romania. Viral RNA extraction was performed automated on the Maxwell 48 RSC Extraction System (Promega, USA) using the Maxwell RSC Viral Total Nucleic Acid Purification kit (Promega, USA). After RNA extraction, the samples were amplified on a 7500 real-time PCR (Applied Biosystems, USA) using the genesig® Real-Time PCR Assay (Primer Design, UK). RESULTS: The molecular testing showed a cycle threshold value of 23.4 (1.2 x 106 copies/mL), indicating increased viral loads, which correlated with the laboratory analysis results, especially with D-dimer levels. CONCLUSIONS: In cases of coagulopathy of SARS-CoV-2, patients in hospitals should be monitored closely for thrombosis development. Thus D-dimer can be used as prognostic marker in monitoring the evolution of SARS-CoV-2 infected patients.

Circulating Cell-Free Plasma DNA in Noninvasive Prenatal Paternity Testing with Short Tandem Repeats (STRs).

BACKGROUND: Paternity relationship can be established using STR markers in a minimally invasive manner during the prenatal period in the early weeks of pregnancy or in advanced pregnancy using circulating cell-free DNA (ccf DNA) drawn from the mother. The aim of our presentation is to demonstrate the advantages of ccf plasma DNA in establishing the paternity of an unborn child. Between mother and the alleged father (AF) of the fetus, an avuncular relationship as uncle-niece exists. METHODS: As biological samples, saliva was collected with buccal swabs from the mother and AF. For the fetus, we separated plasma from drawn blood from the mother, and further, we isolated ccf DNA from the mother's plasma sample. The DNA samples were quantified on a 7500 ABI Real-Time PCR using Investigator Quantiplex Pro Kit (Qiagen, Germany). Genotyping of the DNA samples was performed on a ProFlex PCR System (Thermo Scientific, USA) using the multiplex STR markers from Global Filer PCR Amplification Kit (Thermo Scientific, USA). Further, PCR products were run on capillary electrophoresis on an ABI 3500 Genetic Analyzer (Applied Biosystems, USA). RESULTS: The AF was confirmed as the biological father of the child, with a probability of paternity (PP) = 99.99999% and a cumulative paternity index (CPI) = 8.300 x 103. CONCLUSIONS: In the case of advanced pregnancies from sexual assaults or incestuous relationships, the use of ccf DNA to establish the genetic profile of the fetus represents an advantage for establishing the paternity relationship between the fetus and AF. The method proves its efficiency as it has the advantage of speed of probation through forensic genetic expertise.

Detection of Mutations in Short Tandem Repeats (STRs) Loci in Paternity Testing in Romanian Population.

BACKGROUND: In forensic genetics, mutation analysis for different short tandem repeat (STR) loci is important for paternity and maternity testing. The aim of this study is determining the most frequent loci with mutations in a population of 743 individuals in western Romania in 246 kinship cases. These include 240 paternity and 6 maternity tests analyzed at the Laboratory of Forensic Genetics, Victor Babes University of Medicine and Pharmacy, Timisoara, Romania. The study was conducted between January 1, 2017, to January 1, 2020. The study aims to analyze the mutation rates for 15 autosomal markers used in this type of testing. The following loci were included in our study: D3S1358, D8S1179, D18S51, D21S11, FGA, TH01, vWA, CSF1PO, D7S820, D13S317, D16S539, D2S1338, D19S433, TPOX, D5S818. METHODS: For the reference samples, we used saliva collected on buccal swabs from all individuals. Salivary DNA was quantified on the 7500 real-time PCR equipment (Thermo Scientific, USA). Further, amplification of the DNA samples was performed on a ProFlex PCR System (Thermo Scientific, USA) using Identifiler Plus PCR Am-plification kit (Thermo Scientific, USA). Fragment analysis was performed on the 3500 Genetic Analyzer (Thermo Scientific, USA). The genetic profiles were generated by GeneMapper ID-X software version 1.4 (Thermo Scientific, USA). RESULTS: The mutation events in paternity testing were observed in 10 out of the 15 analyzed loci: D21S11, D18S51, D16S539, D8S1179, FGA, D2S441, D19S433, D2S1338, D3S1358, D5S818 and vWA. Paternal mutations were more frequent (63%) than maternal mutations (37%). CONCLUSIONS: The results confirm that the mutation rate in paternity tests are more frequent during paternal meiosis compared to maternal.

Advantages of Chromosome X-STRs Markers in Solving a Father-Daughter Paternity Case with one Mismatch on SE33 Locus.

BACKGROUND: Genetic markers are routinely used in human identification of paternity, maternity, and kinship cases. We describe a DNA paternity case with one mismatch on SE33 locus between the alleged father (AF) and the child (daughter). Because there was a father-daughter relationship to solve this case we used chromosome X-STRs markers too. METHODS: As reference samples we used saliva collected from inside the cheek of each person using buccal swabs (Copan, Italy). The DNA samples were quantified on a 7500 ABI real-time PCR using the Investigator Quantiplex Pro Kit (Qiagen, Germany). Salivary DNA samples were amplified on a ProFlex PCR System (ThermoFischer, USA) using the multiplex STR markers from the AmpF/STR® NGM Select PCR Amplification Kit (Thermo-Fischer, USA) and Investigator® Argus X-12 QS kit markers (Qiagen, Germany). PCR products were run on capillary electrophoresis on an ABI 3500 Genetic Analyzer (ThermoFischer, USA). RESULTS: The AF was excluded from paternity on STRs markers due to one mismatch on SE33 locus. To confirm or exclude the paternity, we used the chromosome X-STRs markers, obtaining a perfect match between the AF and his daughter. CONCLUSIONS: In paternity testing, where one or two mismatches are present between the child (daughter) and the AF on different loci on STR markers, the use of chromosome X-STRs is needed for the confirmation or exclusion of paternity.

Co-morbidities in the multiple victims of the silent killer in carbon monoxide poisoning.

Carbon monoxide (CO) remains an insidious and silent killer due to its physical and chemical properties; its lethal effects are encountered in cases of household accidents, occupational hazards or suicide. Deaths due to CO poisoning were studied retrospectively in the period 2000-2018 at the Institute of Forensic Medicine, Timisoara, Romania. These cases represent 1.75% of all the autopsies and 0.63% of all violent deaths. There have been cases of single deaths and cases with multiple victims - concomitant deaths. The analysis of lethal CO intoxication cases that occurred in different circumstances (incomplete burning with CO accumulation, fires - associated with burns, death in the fountain - due to fossil fuel pump failure, suicide due to exhaust gases) was based on the examination of 298 autopsy files. In this type of poisoning, the forensic examination of the body is marked by the non-specific character of most of the macroscopic and microscopic changes. Although inconstant, these types of changes (e.g., red discoloration of livor mortis) raise the suspicion of death by CO poisoning; the essential contribution to establishing cause of death resides in the determination of carboxyhemoglobin (COHb) concentration by spectroscopy. In all cases, the cerebral and cardio-pulmonary modification and their contribution to the cause of death were studied. Co-morbidities interfere with the cause of death in cases with average COHb concentrations, in the 20-50% range, where CO blood levels alone are not reason enough to explain the onset of death.

A Rare Variant of the Great Cardiac Vein Draining Directly into the Superior Vena Cava.

OBJECTIVE: A routine dissection of the cadaver of a 67-year-old man revealed a very rare morphological variant of the great cardiac vein (GCV). PRESENTATION: The vein originated in the upper third of the anterior interventricular sulcus, crossed the anterior interventricular artery superficially, ran beneath the circumflex artery, crossed the transverse pericardial sinus, and drained directly into the superior vena cava. CONCLUSION: This variant of the GCV is interesting due to its rarity. It is important to know about it for procedures that require venous access such as coronary surgery requiring retrograde cardioplegia, surgical ablation of aberrant conducting pathways, pacemaker insertion, and valve surgery.

The multi-factorial determinism of forensic expertise regarding sentence interruption on medical grounds and decision.

The topic proposed by this research is the complex determinism or requesting forensic expertise for the purpose of postponing or interrupting a sentence in medical ground. Within the topic, it was necessary to perform a complex analysis on several aspects of inmate life, from the point of view of their rights and especially the right to medical assistance. We have included aspects related to the evolution of the legal framework and current regulations, to the status of medical assistance in the penitentiary environment (illustrating realities related to morbidity and mortality) and to the doctor-patient relationship during a forensic expertise. To this end, this paper aims to reveal the realities of how inmates are informed about the framework in which the institution of forensic expertise operates, as well as to analyse the inmates' perception on the accessibility and usability of this type of expertise. At the same time, we aimed to identify cases which abusively exceed the boundaries of requesting sentence interruption on medical grounds and the causes of these situations. In the first part, a retrospective statistical study was performed in mortality within the penitentiary population in the area included in the study. The characteristics particular to the doctor-inmate patient relationship were also explored.

Chimerism Monitoring by Short Tandem Repeat (STR) Markers in Allogeneic Stem Cell Transplantation.

BACKGROUND: Allogeneic hematopoietic stem cell (allo-HSC) transplantation is used in the treatment of malignant hematological diseases. An important tool in monitoring post-transplantation evolution is represented by the percentage of donor's blood cells found in recipient's blood, known as chimersim. This is useful in predicting the graft rejection and the risk of disease relapse. In this study, we present the importance of multiplex STR markers in chimerism monitoring of a 8 year old girl diagnosed with acute lymphoblastic leukemia (ALL). METHODS: In the pre-transplant stage, saliva on buccal swabs and blood samples in EDTA were collected from the donor and recipient and used as reference samples. The DNA extraction from saliva and blood samples was done using the Pure Link Genomic DNA kit (Invitrogen, USA). For the DNA quantification, the Quantifiler Human DNA kit (Applied Biosystems, USA) was used on an ABI 7500 Real-time PCR system (Applied Biosystems, USA). Amplification of the STR markers was performed using the AmpFLSTR NGM SElect kit (Applied Biosystems, USA) on a ProFlex PCR System. The PCR products were separated and detected on an ABI 3500 Genetic Analyzer (Applied Biosytems, USA). RESULTS: One month post-transplantation of HSC, a mixed chimerism (MC) containing 38% of donor's cells was obtained from a bone marrow aspiration sample. On the 45th day, a new transplantation was performed. On the 15th day after 2nd transplantation, a MC with 91% donor's cells was obtained. On the 21st day after the 2nd transplantation, a complete chimerism (CC) with 100% donor's cells was obtained. CONCLUSIONS: Chimerism monitoring is useful in identifying those patients in risk for relapse or graft rejection.

A Single Step Mutation at D3S1358 Locus in a DNA Paternity Testing with 2 Alleged Fathers.

BACKGROUND: Genetic information is used very frequently in human identification in civil or judicial cases. Establishing the kinship relationship between a child and his biological father involves many ethical facts. We describe a DNA paternity case with two alleged fathers and an inconsistency between alleged father-2 and the child at D3S1358 locus. METHODS: As biological samples we used saliva collected from inside the cheek of each person using buccal swabs (Copan, Italy). We collected the biological samples from each of person after each person gave the consent. In order to find the concentration of salivary DNA, the DNA samples were quantified by 7500 ABI Real-time PCR using the Quantifiler Human DNA kit (Applied Biosystems, USA). The next step was the amplification of the Salivary DNA samples by polymerase chain reaction (PCR). It was performed on a ProFlex PCR System (Applied Biosystems, USA) using the multiplex STR markers from the AmpFlSTR® Identifiler Plus Amplification Kit (Applied Biosystems, USA). After amplification, the PCR products were run on capillary electrophoresis on an ABI 3500 Genetic Analyzer (Applied Biosystems, USA). RESULTS: AF-1 was excluded as biological father. The DNA profiles of AF-2 and the child had one mismatch at D3S1358 locus. Further, we amplified the Y-STR markers to confirm the mutation, obtaining a perfect match between the 2 persons. CONCLUSIONS: In paternity testing, where one or two inconsistencies are present between the child and the alleged father on autosomal STR markers, the use of haploid markers X-STR or Y-STRs is needed for the confirmation or exclusion of paternity.