A reference database of forensic autosomal and gonosomal STR markers in the Tigray population of Ethiopia.

Allele frequencies of 21 autosomal STR markers (AmpF/STR GlobalFiler) and haplotype frequencies of 27 Y- and 12 X-STR markers (AmpF/STR YFiler Plus and Investigator Argus X-12, respectively) were investigated in the Tigray population of Ethiopia, representing the main population group in the Tigray regional state of Ethiopia and neighboring Eritrea. For autosomal STR allele frequencies, the average random match probability in the Tigray sample was 2.1 × 10-27. The average locus by locus FST distance calculated comparing autosomal STR allele frequencies from Tigray and from a broad regional reference dataset currently available for the Horn of Africa was 0.003. The Tigray male sample displayed high Y-STR diversity, with complete individualization of haplotypes using the AmpF/STR YFiler Plus panel. Analysis of molecular variance did not detect significant heterogeneity between Y-STR haplotypes observed in the present study and those previously reported in the literature for other Tigray population samples from Ethiopia and Eritrea. Study of the X-STR landscape in Tigray evidenced several distinctive features including: the molecular characterization of a novel null allele at locus DXS10146 with frequency > 1%; allele dependency between loci within linkage groups I and III; significant differences in haplotype distribution compared to other Horn of Africa populations, that should be taken into account in kinship analysis. The collected data can be used as a reference STR database by local forensic genetics services and in genetic identification procedures of victims of human trafficking in the Mediterranean Sea, which frequently involve individuals originating from the Horn of Africa.

A Ge.F.I. - ISFG European collaborative study on DNA identification of Cannabis sativa samples using a 13-locus multiplex STR method.

Cannabis sativa is the most used controlled substance in Europe. With the advent of new and less restrictive European laws on cannabis sale for recreational use (including in Italy), an increase in indoor cannabis crops were observed. This increase was possible due to the availability of cannabis seeds through the internet market. Genetic identification of cannabis can link seizures and if in possession then might aid in an investigation. A 13-locus multiplex STR method was previously developed and validated by Houston et al. A collaborative exercise was organized by the Italian Forensic Geneticists - International Society of Forensic Genetics (Ge.F.I. - ISFG) Working Group with the aim to test the reproducibility, reliability and robustness of this multiplex cannabis STR kit. Twenty-one laboratories from three European countries participated in the collaborative exercise and were asked to perform STR typing of two cannabis samples. Cannabis DNA samples and the multiplex STR kit were provided by the University of Barcelona and Sam Houston State University. Different platforms for PCR amplification, capillary electrophoresis (CE) and genotyping software were selected at the discretion of the participating laboratories. Although the participating laboratories used different PCR equipment, CE platforms and genotyping software, concordant results were obtained from the majority of the samples. The overall genotyping success ratio was 96%. Only minor artifacts were observed. The mean peak height ratio was estimated to be 76.3% and 78.1% for sample 1 and sample 2, respectively. The lowest amount of -1 / + 1 stutter percentage produced, when the height of the parent allele was higher than 8000 RFU, resulted to be less than 10% of the parent allele height. Few common issues were observed such as a minor peak imbalance in some heterozygous loci, some artifact peaks and few instances of allelic drop-out. The results of this collaborative exercise demonstrated the robustness and applicability of the 13-locus system for cannabis DNA profiling for forensic purposes.

Characterization of ancestry informative markers in the Tigray population of Ethiopia: A contribution to the identification process of dead migrants in the Mediterranean Sea.

Determination of bio-geographical ancestry by means of DNA ancestry informative markers (AIMs) can contribute to the identification of human remains in missing person cases and mass disasters. While the presence of Eastern Africans among the migrant victims of trafficking accidents in the Mediterranean Sea is often suspected, few studies have addressed the ability of autosomal AIM panels in current use in forensic laboratories to provide differentiation of populations within the African continent. In this study, two assays consisting of 46 AIM-Indels and 31 AIM-SNPs were typed in a Tigray population sample from Northern Ethiopia. STRUCTURE analysis showed that the Tigray population is characterized by a strong (∼50 %) non-African genetic component shared with European and Middle Eastern populations. The intermediate position of the Tigray sample between sub-Saharan African and European / Middle Eastern reference population samples was confirmed by principal component analysis. Both AIM panels provided effective differentiation between Tigray and sub-Saharan African populations. Classification accuracy of other populations involved in the current Mediterranean migrant crisis, like South Asians, was superior with the AIM-SNP panel compared to the AIM-Indel panel. Misclassification of Middle Eastern samples as Tigray was frequent with both AIM-indel (∼30 % misclassified) and AIM-SNPs (∼20 %). However, with AIM-SNPs, error rates were reduced to acceptable levels by applying cautionary minimum thresholds to assignment likelihoods. Establishment of an Eastern African reference database of AIMs that can be genotyped by means of low cost, small-scale assays compatible with capillary electrophoresis, sets a balance between the need for ancestry inference tools and the budget limitations faced by Italian laboratories engaged in the humanitarian identification of dead migrants recovered from the Mediterranean Sea.

Subtyping of Y-chromosomal haplogroup E-M78 (E1b1b1a) by SNP assay and its forensic application.

The continual discovery of new single-nucleotide polymorphisms (SNPs) has led to an increased resolution of the Y chromosome phylogeny. Some of these Y-SNPs have shown to be restricted to small geographical regions and therefore may prove useful in the forensic field as tools for the prediction of population of origin of unknown casework samples. Here, we describe a system for the molecular dissection of haplogroup E-M78 (E1b1b1a), consisting of multiplex polymerase chain reaction and minisequencing of M78 and nine population-informative Y-SNPs (M148, M224, V12, V13, V19, V22, V27, V32, V65) in a single reaction. Sensitivity and admixture studies demonstrated that the SNP protocol allows robust genotyping from as little as 50 pg of male DNA, even in the presence of 500-fold amounts of female DNA. In order to evaluate the suitability of E1b1b1a, subhaplogrouping for population-of-origin prediction, the distribution of E-M78 and its derived variants was determined in an Italian population sample (n = 326).

Allele and haplotype diversity of 12 X-STRs in Sardinia.

The analysis of clusters of tightly linked X-chromosome short tandem repeat (STR) markers can assist the interpretation of complex kinship cases. However, when linkage disequilibrium (LD) is present in the population of origin of tested individuals, haplotype rather than allele frequencies should be used in likelihood calculations. The diversity of twelve X-STRs arranged in four linkage groups (I: DXS10148-DXS10135-DXS8378; II: DXS7132-DXS10079-DXS10074; III: DXS10103-HPRTB-DXS10101; IV: DXS10146-DXS10134-DXS7423) was tested in a Sardinian population sample (n=516) including three open populations from the Northern, Central and Southern part of the island, and three isolates (Benetutti, Desulo, Carloforte). Evidence of LD was detected in Sardinia within each linkage group. Significant differences in haplotype and allele frequency distribution of X-STR markers was seen between isolates and open populations, which on the contrary appeared highly homogeneous. The percentage of Sardinian haplotypes previously unobserved in a similar dataset compiled for the Italian population was: 76.3% (linkage group I), 61.3% (linkage group II), 54.1% (linkage group III), 58.9% (linkage group IV). Significant pairwise genetic differences were seen between mainland Italy, the three Sardinian isolates, and the open population of Southern Sardinia. The study confirms the presence of high levels and complex patterns of LD along the X chromosome in Sardinia, and provides population-specific haplotype data for biostatistical evaluation in kinship testing.

Investigation of extended Y chromosome STR haplotypes in Sardinia.

Y-chromosomal variation of selected single nucleotide polymorphisms (SNPs) and 32 short tandem repeat (STR) loci was evaluated in Sardinia in three open population groups (Northern Sardinia, n=40; Central Sardinia, n=56; Southern Sardinia, n=91) and three isolates (Desulo, n=34; Benetutti, n=45, Carloforte, n=42). The tested Y-STRs consisted of Yfiler® Plus markers and the seven rapidly mutating (RM) loci not included in the YFiler® Plus kit (DYF399S1, DYF403S1ab, DYF404S1, DYS526ab, DYS547, DYS612, and DYS626). As expected, inclusion of additional Y-STR loci increased haplotype diversity (h), though complete differentiation of male lineages was impossible even by means of RM Y-STRs (h=0.99997). Analysis of molecular variance indicated that the three open populations were fairly homogeneous, whereas signs of genetic heterogeneity could be detected when the three isolates were also included in the analysis. Multidimensional scaling analysis showed that, even for extended haplotypes including RM Y-STR markers, Sardinians were clearly differentiated from populations of the Italian peninsula and Sicily. The only exception was represented by the Carloforte sample that, in accordance with its peculiar population history, clustered with Northern/Central Italian populations. The introduction of extended forensic Y-STR panels, including highly variable RM Y-STR markers, is expected to reduce the impact of population structure on haplotype frequency estimations. However, our results show that the availability of geographically detailed reference databases is still important for the assessment of the evidential value of a Y-haplotype match.

Y-chromosomal STR haplotypes in Sicily.

Eight Y-chromosomal short tandem repeats (STRs)-DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393 and DYS385-were typed in a population sample (n=255) of unrelated Sicilian males from nine different towns on the main island and from the island of Pantelleria.

A molecular exploration of human DNA/RNA co-extracted from the palmar surface of the hands and fingers.

"Touch DNA" refers to the DNA that is left behind when a person touches or comes into contact with an item. However, the source of touch DNA is still debated and the large variability in DNA yield from casework samples suggests that, besides skin, various body fluids can be transferred through contact. Another important issue concerning touch DNA is the possible occurrence of secondary transfer, but the data published in the literature in relation to the background levels of foreign DNA present on the hand surfaces of the general population are very limited. As the present study aimed at better understanding the nature and characteristics of touch DNA, samples were collected from the palmar surface of the hands and fingers ("PHF" samples) of 30 male and 30 female donors by tape-lifting/swabbing and subjected to DNA/RNA co-extraction. Multiplex mRNA profiling showed that cellular material different from skin could be observed in 15% of the PHF samples. The total amount of DNA recovered from these samples (median 5.1 ng) was significantly higher than that obtained from samples containing skin cells only (median 1.6 ng). The integrity of the DNA isolated from the donors' hands and fingers as well as the prevalence of DNA mixtures were evaluated by STR typing and compared with reference STR profiles from buccal swabs. DNA integrity appeared significantly higher in the male rather than in the female subsample, as the average percentage of the donors' alleles effectively detected in PHF profiles was 75.1% and 60.1%, respectively. The prevalence of mixtures with a foreign DNA contribution ≥20% was 19.2% (30.0% in the female PHF samples and 8.3% in the male PHF samples). The obtained results support the hypothesis that transfer of cellular material different from skin may underlie the occasional recovery of quality STR profiles from handled items. These results also suggest that gender may represent an important factor influencing the propensity of individuals to carry and transfer DNA through hand contact, possibly because of the differences in personal and hygiene habits between males and females.

Evaluation of DNA typing as a positive identification method for soft and hard tissues immersed in strong acids.

Identification of human remains can be hindered by several factors (e.g., traumatic mutilation, carbonization or decomposition). Moreover, in some criminal cases, offenders may purposely adopt various expedients to thwart the victim's identification, including the dissolution of body tissues by the use of corrosive reagents, as repeatedly reported in the past for Mafia-related murders. By means of an animal model, namely porcine samples, we evaluated standard DNA typing as a method for identifying soft (muscle) and hard (bone and teeth) tissues immersed in strong acids (hydrochloric, nitric and sulfuric acid) or in mixtures of acids (aqua regia). Samples were tested at different time intervals, ranging between 2 and 6h (soft tissues) and 2-28 days (hard tissues). It was shown that, in every type of acid, complete degradation of the DNA extracted from soft tissues preceded tissue dissolution and could be observed within 4h of immersion. Conversely, high molecular weight DNA amenable to STR analysis could be isolated from hard tissues as long as cortical bone fragments were still present (28 days for sulfuric acid, 7 days for nitric acid, 2 days for hydrochloric acid and aqua regia), or the integrity of the dental pulp chamber was preserved (7 days, in sulfuric acid only). The results indicate that DNA profiling of acid-treated body parts (in particular, cortical bone) is still feasible at advanced stages of corrosion, even when the morphological methods used in forensic anthropology and odontology can no longer be applied for identification purposes.

Development of an Italian RM Y-STR haplotype database: Results of the 2013 GEFI collaborative exercise.

Recently introduced rapidly mutating Y-chromosomal short tandem repeat (RM Y-STR) loci, displaying a multiple-fold higher mutation rate relative to any other Y-STRs, including those conventionally used in forensic casework, have been demonstrated to improve the resolution of male lineage differentiation and to allow male relative separation usually impossible with standard Y-STRs. However, large and geographically-detailed frequency haplotype databases are required to estimate the statistical weight of RM Y-STR haplotype matches if observed in forensic casework. With this in mind, the Italian Working Group (GEFI) of the International Society for Forensic Genetics launched a collaborative exercise aimed at generating an Italian quality controlled forensic RM Y-STR haplotype database. Overall 1509 male individuals from 13 regional populations covering northern, central and southern areas of the Italian peninsula plus Sicily were collected, including both "rural" and "urban" samples classified according to population density in the sampling area. A subset of individuals was additionally genotyped for Y-STR loci included in the Yfiler and PowerPlex Y23 (PPY23) systems (75% and 62%, respectively), allowing the comparison of RM and conventional Y-STRs. Considering the whole set of 13 RM Y-STRs, 1501 unique haplotypes were observed among the 1509 sampled Italian men with a haplotype diversity of 0.999996, largely superior to Yfiler and PPY23 with 0.999914 and 0.999950, respectively. AMOVA indicated that 99.996% of the haplotype variation was within populations, confirming that genetic-geographic structure is almost undetected by RM Y-STRs. Haplotype sharing among regional Italian populations was not observed at all with the complete set of 13 RM Y-STRs. Haplotype sharing within Italian populations was very rare (0.27% non-unique haplotypes), and lower in urban (0.22%) than rural (0.29%) areas. Additionally, 422 father-son pairs were investigated, and 20.1% of them could be discriminated by the whole set of 13 RM Y-STRs, which was very close to the theoretically expected estimate of 19.5% given the mutation rates of the markers used. Results obtained from a high-coverage Italian haplotype dataset confirm on the regional scale the exceptional ability of RM Y-STRs to resolve male lineages previously observed globally, and attest the unsurpassed value of RM Y-STRs for male-relative differentiation purposes.

Granulomatous renal disease in a patient with common variable immunodeficiency.

Common variable immunodeficiency (CVID), the most common cause of primary hypogammaglobulinemia, is characterized by a decreased serum immunoglobulin level, recurrent infections, and the occurrence of various autoimmune diseases. Granulomatous disease has been reported previously in several patients with CVID, with granuloma occurring in the lymph nodes, spleen, liver, central nervous system, and bone marrow. We report the first published case of renal granulomatous disease in a CVID patient presenting with subacute renal failure. Renal function partially recovered after corticosteroid treatment and intravenous immunoglobulin infusions. The pathogenesis of granulomatous disease in CVID is unclear but may involve monocyte and T-cell abnormalities.

Y-chromosomal STR haplotypes in an Albanian population sample.

Eight Y-chromosomal short tandem repeats (STRs), DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393 and DYS385, were typed in a population sample (n=101) of first-generation Albanian immigrants living in Italy.

Y-chromosomal STR haplotypes in a population sample from continental Greece, and the islands of Crete and Chios.

Eight Y-chromosomal short tandem repeats (STRs)--DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, and DYS385--were typed in a population sample (n = 113) of unrelated males from seven different regions of Greece (Macedonia, Thessaly, Epirus, Central Greece, Peloponnese, Crete Island, and Chios Island).

LDLR, GYPA, HBGG, D7S8 and GC allele and genotype frequencies in the northwest Italian population.

Allele and genotype frequencies for five PCR-based DNA markers (LDLR, GYPA, HBGG, D7S8 and GC) were determined in 100 unrelated individuals from Piedmont (Northwest Italy). All five Ioci met Hardy-Weinberg expectations in the sampled population. The combined PD and CE were, respectively, 0.995 and 0.697. Frequencies obtained were compared with other previously published data on Caucasian populations with no significant differences. The genetic data from this study, in addition to those already collected by other groups, contribute to the expansion of the Italian DNA database suitable for forensic casework and paternity testing.

[Biological monitoring of chronic hemodialysis]. / Suivi biologique de l'hémodialyse chronique.

Patients suffering from acute renal failure must undergo dialysis to substitute for the kidney's excretory function. Dialysis is a means of exchange between two solutions: blood and a liquid known as dialysate, across a semi-permeable membrane. This membrane permits the passage of water and aqueous solutions of low molecular weight but not that of the aqueous solutions of high molecular weight such as proteins. Dialysed patients are biologically monitored to prevent the various complications arising from blood dialysis, to check the efficacy of the treatment as well as to reduce the risk of morbidity and mortality. Because of the multiple and complex nature of renal functions, biological monitoring of a dialysis patients presents many, often inter-related facets. In addition to the complications inherent in dialysis itself, the dialysed patient is exposed to other causes of mortality or morbidity such as nutritional, hematologic, cardiovascular and infectious problems. Clinicians is confronted by other complications, albeit less common and easier to control, in the form of osteoarticular troubles or even aluminium poisoning. Lastly, biological assessment of the efficacy of purifying processing depends principally on calculating the dose of dialysate and the ureic index of subtraction which, in turn, requires the measurement of uremia.

[Indications and preparations for kidney dialysis]. / Indication et préparation au traitement de suppléance rénale.

Every patient with end-stage renal failure, at any age and whatever the type of renal disease, is a legitimate candidate to maintenance dialysis. Contraindications are infrequent and based purely on medical considerations, such as profound and irremediable alteration of physical and/or mental condition. In patients regularly managed dialysis is decided electively on the basis of laboratory criteria in the absence of clinical uremic manifestations other than fatigue, anorexia or nausea. The most widely accepted criterion is a level of creatinine clearance estimated by the Cockcroft-Gault formula between 7 and 10 mL/min/1.73 m2. Psychological preparation of the patient to dialysis is essential and should not be delayed until the advanced stage. Medical preparation involves prophylactic vaccination against virus B hepatitis and creation of a native arteriovenous fistula when hemodialysis is the scheduled option. Every patient should receive in time clear and complete information on the various technical methods of dialysis, in order to allow him an informed choice.

Approaches in injections for radicular pain: the transforaminal, epidural and transfacet approaches.

Spinal injections must be carried out adhering to very strict conditions. However, these procedures have almost come to be seen as everyday and may be practised under quite questionable conditions. The recent reports of new and extremely serious neurological complications have changed the attitudes of those making referrals as well as the attitudes of the interventional radiologists carrying out these procedures. The range of indications for transforaminal injections has shrunk in favour of epidural injections. Where the transforaminal approach is still used, the needle must be positioned extremely accurately. A prior radioopaque contrast medium injection is essential from a safety perspective. The transforaminal epidural injection via the transfacet approach looks to be a promising alternative that is strictly avascular.