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This article reports the determination of phenobarbital in the mummified body of a 56-year-old man found completely mummified 10 years after his death. When alive, he was being treated for epilepsy with phenobarbital, and the recent analyses, performed with both immunochemical techniques and gas chromatography with mass spectrometry (GC-MS), have revealed the presence of this substance in various tissues: the mean content of barbiturate in the mummified liver tissue was 93 μg/g, 216 μg/g in the heart, 17 μg/g in the lungs, 12 μg/g in muscles, and 31 μg/g in the skin. Preliminary screening tests with immunochemical techniques to evaluate the presence of other drugs were also performed. The sample resulted negative for all substances tested. Phenobarbital can be identified and quantified thanks to its excellent chemical stability and a hypothesis of what the concentrations in the fresh tissue could have been has also been reported.

Many X-chromosome short tandem repeats (X-STRs) have been validated for forensic use even if further studies are needed on allele frequencies and mutation rates to evaluate the extent of polymorphism in different populations and to establish reference databases useful for forensic applications and for anthropological studies. A single multiplex reaction of seven X-STRs, which includes the DXS6789, HUMARA, DXS10011, DXS7423, HPRTB, DXS6807, DXS101 loci, is presented and their allele frequency distribution in a large population sample including 556 subjects (268 females and 288 males) analysed by five forensic laboratories of Central and Northern Italy is shown. Our results demonstrate the feasibility of a single amplification/detection reaction involving seven markers of the X chromosome, which can be fruitfully used in complex kinship analysis.

Background: Phylogenetic and genetic analyses have proven a valuable tool to infer epidemiological links between human immunodeficiency virus type-1 (HIV-1) isolates. These methods were applied in the present report for studying the genetic relatedness of the viral strains involved in two episodes of suspected HIV-1 transmission. Objectives: Provide any evidence that may help establish or refute the transmission link. Study design: In the first case, a leukemic patient became HIV-1 positive following the transfusion of platelets from a donor who was subsequently found to have tested false HIV-seronegative and to be sexual partner to an infected woman. In the second, a wife claimed to have acquired the infection from her husband who had concealed his infected status. Results and conclusions: The viral pairs detected in each of the suspected transmission cases exhibited common amino acid signatures and low genetic distances and segregated together in phylogenetic trees, thus showing a level of genetic relatedness similar to reference pairs known with certainty to be epidemiologically linked. These findings corroborated the existence of a direct transmission link in both the episodes with a high level of confidence.

We report on 34 subjects from a small Italian town distributed among 13 different surnames. Their genealogies have been reconstructed to the most recent common ancestor, going back in a case to the 15th century, and their haplotypes at 12 Y-chromosome STR has been determined. Six surnames (14 subjects) did not show variation at any locus among isonymous individuals, whereas seven surnames (20 subjects) showed single-step differences at single loci (two in DYS391 and DYS390, the others in DYS385, DYS392 and DYS393); in addition, two subjects of two different surnames showed 8 and 9 locus differences, respectively.

The role of DNA damage in PCR processivity/fidelity is a relevant topic in molecular investigation of aged/forensic samples. In order to reproduce one of the most common lesions occurring in postmortem tissues, a new protocol based on aqueous hydrolysis of the DNA was developed in vitro. Twenty-five forensic laboratories were then provided with 3.0 μg of a trial sample (TS) exhibiting, in mean, the loss of 1 base of 20, and a molecular weight below 300 bp. Each participating laboratory could freely choose any combination of methods, leading to the quantification and to the definition of the STR profile of the TS, through the documentation of each step of the analytical approaches selected. The results of the TS quantification by qPCR showed significant differences in the amount of DNA recorded by the participating laboratories using different commercial kits. These data show that only DNA quantification "relative" to the used kit (probe) is possible, being the "absolute" amount of DNA inversely related to the length of the target region (r(2) = 0.891). In addition, our results indicate that the absence of a shared stable and certified reference quantitative standard is also likely involved. STR profiling was carried out selecting five different commercial kits and amplifying the TS for a total number of 212 multiplex PCRs, thus representing an interesting overview of the different analytical protocols used by the participating laboratories. Nine laboratories decided to characterize the TS using a single kit, with a number of amplifications varying from 2 to 12, obtaining only partial STR profiles. Most of the participants determined partial or full profiles using a combination of two or more kits, and a number of amplifications varying from 2 to 27. The performance of each laboratory was described in terms of number of correctly characterized loci, dropped-out markers, unreliable genotypes, and incorrect results. The incidence of unreliable and incorrect genotypes was found to be higher for participants carrying out a limited number of amplifications, insufficient to define the correct genotypes from damaged DNA samples such as the TS. Finally, from a dataset containing about 4500 amplicons, the frequency of PCR artifacts (allele dropout, allele drop-in, and allelic imbalance) was calculated for each kit showing that the new chemistry of the kits is not able to overcome the concern of template-related factors. The results of this collaborative exercise emphasize the advantages of using a standardized degraded DNA sample in the definition of which analytical parameters are critical for the outcome of the STR profiles.

The 2011 collaborative exercise of the ISFG Italian Working Group GeFI was aimed at validating the five ENFSI/EDNAP miniSTR loci D1S1656, D2S441, D10S1248, D12S391 and D22S1045. The protocol required to type at least 50 multilocus profiles from locally resident individuals and two blind bloodstains in duplicate (i.e., using at least two different commercial kits), and to send the electropherograms to the Organizing Committee. Nineteen laboratories distributed across Italy participated, collecting a total of 960 samples. Full concordance was found for the five new miniSTRs as observed from the comparison of 13,150 alleles. The inspection of the electropherograms allowed the identification of a very limited number of mistypings in the miniSTR genotypes thus contributing to the establishment of an high quality Italian database of frequencies.

Haplotype data of 1176 Italian males from 10 regions were obtained as a part of a collaborative validation exercise. Individual data are available at http://www.gefi-forensicDNA.it .

We collected published and unpublished data from 17 contributing groups participating in the GeFI (Italian Forensic Geneticists). The total number of typed subjects was 1114 males and 777 females, coming from 11 regions of North, Centre, and South Italy, and Sardinia. Individual's multilocus genotypes included 4–12 loci. The total number of typed markers was 29, scattered along the X-chromosome genetic map in several clusters; the most used marker was DXS7423 (2429 gene copies); the mean number of subjects typed per marker was 336 for males and 208 for females. Data are available online.

Within an Italian collaborative exercise on the extended haplotype of the Y-chromosome, 1288 subjects were typed by the AmpFlSTR YFiler Amplification Kit (AB Applied Biosystems) and other 526 were typed by the PowerPlex Y ® System (Promega). The sampling scheme included either a "regional" or a "local" recruitment, the first referring to individuals born in the region of the participating lab, the second referring to individuals coming from small villages. Total sample sizes were N =954 and 860, respectively. A significant decrease of haplotype diversity was found in the local samples. The results may be of interest in forensic applications of the Y-chromosome.

A sample of 1070 individuals was collected by 12 collaborating forensic laboratories from Central and Northern Italy, and typed by the so-called "NC01" triplex, including the three loci D10S1248, D14S1434, and D22S1045. Heterozygosities were 0.771, 0.717, and 0.720, respectively.

BACKGROUND: The usual set of autosomal markers (A-STRs) available in commercial kits is often insufficient to discriminate between close relationships when only two subjects are available for analysis. X-chromosome markers (X-STRs) provide higher statistical power in special cases. STUDY DESIGN AND METHODS: Formulas are derived for the probabilities of all possible genotype pairs for X-STRs of any sex combination for seven common relationships. The power of exclusion (PE) of X-STRs in parentage analysis is compared with that of A-STRs of equivalent distribution of allele frequency. RESULTS: Seventy-three equations were obtained, from which the likelihood ratio of any two alternative hypotheses about the relationship between two individuals can be obtained by division and simplification. For father-daughter and mother-son duos, the PE of X-STRs is almost twice the corresponding value of A-STRs for moderately low values of heterozygosity (0.6-0.75); for alleged pairs of sisters and pairs of half-sisters the PE is equivalent to that of A-STRs in parent-child duos. Considering four real unlinked X-STRs, the cumulative PE for father-daughter and mother-son duos was 99 percent, compared with 94 percent if they were autosomal. CONCLUSIONS: X-STRs can substantially increase the discrimination capacity of standard A-STRs in parentage analyses involving pairs of individuals. Up to four unlinked X-STRs may be treated as independent loci. When linked loci are included, computer programs that calculate pedigree likelihoods can be used.

A deficiency case of inheritance dispute involved two women that were either half sisters or unrelated. Using 16 standard forensic markers, the likelihood ratio (LR) in favour of the first hypothesis was 701.1 (P=0.9986); using four unlinked X-chromosome (chr-X) markers (DXS101, HPRTB, STRX1, DXS8377), the LR was 495.8 (P=0.9980). The increase of power in discriminating relationships among females using chr-X markers was investigated by calculating the expected value of the posterior probability that two true half-sisters were half- sisters rather than unrelated for these chr-X marker and autosomal markers with equivalent allele frequency distribution. The mean increase of the posterior probability was included between 19% (HPRTB) and 37% (DXS8377), and was correlated with locus heterozygosity. D 2005 Published by Elsevier B.V.

The inference about the biological relationship between pairs of individuals using genetic markers plays a central role in many areas of human genetics. We investigated the number of markers (M) that are necessary to assign a given proportion of pairs (50%, 60%, 70%, 80%, 90%, 95%, and 99%) to their correct relationship at three predefined probability levels (99%, 99.9%, and 99.99%) against several alternative hypotheses. The following relationships were considered: (1) full sibs (FS), (2) second degree (2D, including half-sibs, grandparent–grandchild and avuncular pairs), (3) first cousins (FC), (4) unrelated individuals (UR).

Allele and genotype frequencies for STR SE33 were obtained for a sample of 419 Italians in view of application in personal identification and paternity. D 2003 Elsevier Science B.V. All rights reserved.

The phylogenetic median-joining network algorithm applied to 111 Y-chromosomes from Tuscany typed at 10 STR loci showed the existence of at least two well-separated clusters of haplotypes. We suggest that they correspond to different haplogroups.

A casework concerning two possible infanticides was solved by a Bayesian approach involving several hypotheses (the mother confessed the crimes following the DNA test). The genetic analysis initially concerned the dead body of an infant found in a rubbish dump and the retained placenta of a woman presenting with massive genital bleeding a year later. After the results of a first genetic test on the two infants, the woman and her current companion were also typed, and the posterior probability that they were parents of both infants was >0.9999, even assuming priors more favorable to the couple.

Six previously characterized STR loci with amplification products smaller than 125 bp have been typed in a sample of 100 unrelated subjects born in Tuscany, Italy, using a protocol involving two triplexes. These loci, called bminiSTRQ, represent promising tools for recovering genetic information from degraded DNA samples when standard loci generate partial profiles. D 2005 Published by Elsevier B.V.

The structural gene of factor B (BF), a component of the alternative complement pathway, is located in the class III region of the major histocompatibility complex (MHC) centromeric to the C2 gene and between the C4 and HLA B genes. Alpern et al (1972) first reported polymorphism of BF, by using immunofixation agarose gel electrophoresis: they identified two common (F and S) and two less common (Fl and S1 = S0.7) alleles, codominant at the same locus. Subsequently, a number of rare variants have been described and a nomenclature has been proposed (Geserick et al 1990). Further isoelectric focusing (IEF) studies revealed the occurrence of BF subtypes (Teng et al 1982; Geserick et al 1983) which could not be demonstrated by standard electrophoresis.

Over the years there has been a continuous increase of the number of genetic polymorphisms which have become available for paternity testing. The range has extended from the red blood cell groups, through serum proteins and erythrocyte enzymes polymorphisms to white cells antigens of the HLA system. With these “classical” markers, the use of several laboratory techniques is needed to achieve a sufficiently high standard of efficiency both in paternity exclusion and inclusion.

In order to achieve an harmonisation among protocols for STRs typing and create a national database, since 1991 the GEFI (Gruppo Ematologi Forensi Italiani) promotes (Pascali 1994, Presciuttini 1994) every two years an inter-laboratory exercise to identify the polymorphic loci more suitable for exchange and comparison of forensic data.

The APO B VNTR locus is located next to the 3′ end of the apolipoprotein B gene and consists of a series of AT rich tandem repeats, each 14–16 bp long (for a review about the genetic relationship between the 3′-VNTR and diallelic apolipoprotein B gene polymorphism, see Renges et al, 1992; for a recent proposal of a molecular model, see Desmarais et al 1993). Its polymorphism was described by Boerwinkle et al (1989) and by Ludwig et al (1989) using the PCR technique. Since then, the APO B has become a genetic marker quite frequently used in forensic practice.

The genetic polymorphism of haptoglobin (HP) has been first observed, through starch gel electrophoresis, by Smithies et al (1955), who demonstrated three common phenotypes: HP 1, HP2-1 and HP2. The HP-α chain, which carries the polymorphism, has subsequently been shown to exist in several additional allelic forms (“subtypes”) - 1S, 1F, 2FS, 2FF and 2SS - giving rise to three different HP1, six different HP2-1 and six different HP2 phenotypes. Only in the last few years, however, methods suitable for routine HP subtyping have been developed.

RFLP (restriction fragment-lenght polymorphism) at the D2S44 locus, detected by the highly polymorphic probe YNH24 (Nakamura et al. 1987), can be used as an efficient tool in stain and paternity testing, as long as reliable fragment size frequencies are available (see van Eede et al. 1991, Gill et al. 1991, Budowle et al. 1991).

The distribution of PLG phenotypes in the population of Tuscany (Central Italy) has been investigated by means of isoelectric focusing followed by immunofixation of desialyzed sera. In a random sample of 383 unrelated healthy blood donors registered at the Hospital of Pisa, three common phenotypes, PLG A, A-B, and B, and two rare variants were found. The allele frequencies calculated in our study were: PLG*A = 0.6749, PLG*B = 0.3225, and PLG*rare = 0.0026. The theoretical exclusion rate in cases of disputed paternity is 17.42%.

Background The traditional exact method for inferring relationships between individuals from genetic data is not easily applicable in all situations that may be encountered in several fields of applied genetics. This study describes an approach that gives affordable results and is easily applicable; it is based on the probabilities that two individuals share 0, 1 or both alleles at a locus identical by state.

The C3 and Bf polymorphisms were studied in 1,000 unrelated Italians. The gene frequencies were calculated and compared with other districts of Italy.

Esterase-D phenotypes were determined in a population sample of Tuscany, Italy. The observed frequencies of the three alleles are: EsD1 = 0.856, EsD2 = 0.143 and EsD3 = 0.001. Studies of the pedigree of the propositus's EsD 3--1 family and mother-child combinations confirm an autosomal co-dominant inheritance.

The thalassaemias, disorders of Hb globin synthesis, and the Hb globin structural variants are hereditary characters with a simple modality of trasmission. Therefore they can be usefully employed in paternity testing.

The distribution of the human red cell esterase D (EsD) “extended” polymorphism in a population sample from Tuscany (Italy) was studied using agarose gel isoelectric focusing. The estimated gene frequencies were: EsD*1 0.864, EsD*2 0.115, EsD*5 0.021. The EsD*5 allele frequency is very similar to those reported for other European populations. The “extension” of the EsD polymorphism may prove to be useful in paternity testing.

Polyacrylamide gel isoelectric focusing (PAGIF) with carrier ampholytes was used for the determination of Tf phenotypes in a sample of 965 unrelated healthy blood donors from Tuscany (Italy). Thirteen rare variants in a heterozygote state were found (four Tf D, seven Tf B, and two rare Tf C subtypes). Among them two apparently new variants, tentatively called Tf C15 and Tf B4, were identified. The rare Tf B0 mutant was also observed.