Samuele Manzoni

Assigning individuals to ethnic groups based on 13 STR loci

Abstract Inferring the ethnic origin of individuals by means of short tandem repeat (STR) profiles has received considerable attention recently. Gene frequency variation among human populations has been extensively documented, and it has been suggested that the differences in allele proportions between ethnic groups could form the basis of an inferential system. We report the use of DNA profiles from 13 STR loci for inferring the ethnic origin of samples of unknown provenance using five populations of immigrants. This preliminary work shows that a population assignment test can be already used in real casework studies.

Determination of endogenous/exogenous γ-hydroxybutyric acid (GHB) in urine

SummaryBackgroundγ-Hydroxybutyric acid (GHB) is an endogenous metabolite and a precursor of the neurotransmitter γ-aminobutyric acid. In some European countries GHB is also used as an anaesthetic agent and in the treatment of alcohol withdrawal. The purpose of the present study was to determine GHB concentrations in urine to enable discrimination between endogenous GHB and exogenous GHB derived from therapeutic administration (Alcover®, Laboratorio farmaceutico C.T.Srl, Sanremo).Methods34 urine samples from 17 alcoholics under treatment for alcohol withdrawal with GHB (group 1) and 12 urine samples from healthy volunteers without GHB intake (group 2) were collected anonymously and analysed by HPLC/MS/MS. The urine concentrations of GHB were determined from two standard curves with the following concentrations: 0.25, 0.50, 1.00, 2.50,5.00 and 10.00 μg/ml, and 5.00, 10.00, 25.00, 50.00 and 100.00 μg/ml.ResultsAcceptable linear regression was obtained for both calibration curves: R2 = 0.9950 and R2 = 0.9994, respectively. Intraday precision and accuracy were calculated as coefficient of variation (%) and bias (n = 10) on three positive control specimens at three different concentrations (0.50, 2.50 and 7.50 μg/ml). The limit of quantification was determined to be 0.50 μg/ml. The median and average concentrations of GHB in group 2 (no GHB intake) were 3.55 and 3.49 μg/ml, respectively, while 52.90% of group 1 (subjects treated with GHB) had urinary GHB concentrations less than the currently used cutoff of 10.00 μg/ml.ConclusionIn the absence of a rapid immunoassay for the detection of GHB in urine, LC/MS/MS analysis was found to be specific and reasonably fast for clinical and forensic toxicology applications. Of the subjects being treated with GHB, 52.90% were found to have a urinary GHB concentration less than 10.00 μg/ml, with GHB concentrations ranging from 2.95 to 8.50 μg/ml. Thus the widely accepted 10.00 μg/ml cut-off could be too high so that there is the risk of false-negative samples.RiassuntoPremesseL’acido γ-idrossibutirrico (GHB) é un metabolita endogeno e precursore del neurotrasmettitore γ-amminobutirrico. In Europa il GHB é anche impiegato come anestetico e come farmaco per il trattamento della dismissione dall’alcol. Lo scopo del presente studio é stato la determinazione della concentrazione urinaria di GHB per poter discriminare i livelli endogeni della molecola da quelli esogeni derivanti dalla somministrazione a scopo terapeutico di GHB (Alcover®, Laboratorio farmaceutico C.T.Srl, Sanremo).Metodi34 campioni di urina di 17 soggetti alcolisti sottoposti al trattamento con GHB per la dismissione dall’alcol (gruppo 1) e 12 campioni di urine di soggetti sani, non bevitori e non in terapia con GHB sono stati raccolti in modo anonimo e sottoposti ad analisi mediante HPLC/MS/MS. La concentrazione urinaria di GHB é stata determinata mediante l’allestimento di due curve di calibrazione: 0,25-0,5-1-2,5-5-10 μg/mL e 5-10-25-50-100 μg/mL.RisultatiEntrambe le curve di calibrazione hanno fornito coeficienti di regressione lineare accettabili: R2 = 0,9950 e R2 = 0,9994 rispettivamente. La precisione intraday e l’accuratezza sono state calcolate mediante la stima del coefficiente di variazione (%) e del bias (n=10) per tre controlli a concentrazioni diverse (0,5 μg/mL, 2,5 μg/mL, 7,5μg/mL). Il limite di quantificazione é stato determinato pari a 0,5 μg/mL. La mediana e la concentrazione media di GHB nel gruppo 2 (non assuntori di GHB esogeno) sono risultate pari a 3,55 e 3,49 μg/mL rispettivamente. Il 52,90% dei soggetti appartenenti al gruppo 1(soggetti trattati con GHB) hanno presentato una concentrazione urinaria inferiore al cut-off utilizzato pari a 10 μg/mL.ConclusioniIn assenza di un test immunochimico rapido per la determinazione di GHB urinario, l’analisi LC/MS/MS puó rappresentare una valida alternativa sia in campo clinico che tossicologico. Il 52,90% dei soggetti trattati con GHB ha presentato una concentrazione urinaria di GHB compresa tra 2,95 e 8,50 μg/mL, ossia inferiore a 10 μg/mL. Si ritiene quindi che un cut-off pari a 10 μg/mL sia troppo alto e che aumenti il rischio di ottenere campioni falsi negativi.

Population data on D16S539, D2S1338 and D19S433 loci in a population sample from Brescia (Italy)

Abstract To establish databases for D16S539, D2S1338 and D19S433 loci for an Italian population sample from Brescia (Northern Italy), 77 unrelated persons were typed. DNA samples were amplified in a multiplex reaction with subsequent automatic detection using capillary electrophoresis. The obtained data are very useful in our practice and they give a contribution to the definition of Italian population short tandem repeat (STR) allelic and genotypic frequencies for the three analysed.