New simple and low-cost methods for periodic checks of Cyclone® Plus Storage Phosphor System

The recent large use of the Cyclone® Plus Storage Phosphor System, especially in European countries, as imaging system for quantification of radiochemical purity of radiopharmaceuticals raised the problem of setting the periodic controls as required by European Legislation. We described simple, low-cost methods for Cyclone® Plus quality controls, which can be useful to evaluate the performance measurement of this imaging system

Pitfalls in the Genetic Identification of Human Remains

DNA technology is an irreplaceable tool for the identification of human remains, but the reliability of the ante mortem reference data remains a serious concern. We present here two cases where misleading conclusions could be achieved by using only the genetic profile of the missing person\u2019s father such as reference sample. Nevertheless, when appropriate reference DNA samples (e.g., the maternal samples) became available, certain identifications were achieved as shown by the probability of maternity (> 99.999%). Thus, all these data together show that extra-pair paternity was found by the way, in both cases. Precautions to avoid misleading conclusions are addressed

Population Structure of Pseudomonas aeruginosa from Five Mediterranean Countries: Evidence for Frequent Recombination and Epidemic Occurrence of CC235

Several studies in recent years have provided evidence that Pseudomonas aeruginosa has a non-clonal population structure punctuated by highly successful epidemic clones or clonal complexes. The role of recombination in the diversification of P. aeruginosa clones has been suggested, but not yet demonstrated using multi-locus sequence typing (MLST). Isolates of P. aeruginosa from five Mediterranean countries (n = 141) were subjected to pulsed-field gel electrophoresis (PFGE), serotyping and PCR targeting the virulence genes exoS and exoU. The occurrence of multi-resistance (≥3 antipseudomonal drugs) was analyzed with disk diffusion according to EUCAST. MLST was performed on a subset of strains (n = 110) most of them had a distinct PFGE variant. MLST data were analyzed with Bionumerics 6.0, using minimal spanning tree (MST) as well as eBURST. Measurement of clonality was assessed by the standardized index of association (IAS). Evidence of recombination was estimated by ClonalFrame as well as SplitsTree4.0. The MST analysis connected 70 sequence types, among which ST235 was by far the most common. ST235 was very frequently associated with the O11 serotype, and frequently displayed multi-resistance and the virulence genotype exoS−/exoU+. ClonalFrame linked several groups previously identified by eBURST and MST, and provided insight to the evolutionary events occurring in the population; the recombination/mutation ratio was found to be 8.4. A Neighbor-Net analysis based on the concatenated sequences revealed a complex network, providing evidence of frequent recombination. The index of association when all the strains were considered indicated a freely recombining population. P. aeruginosa isolates from the Mediterranean countries display an epidemic population structure, particularly dominated by ST235-O11, which has earlier also been coupled to the spread of ß-lactamases in many countries

DNA extraction from blood and forensic samples

Genetic analyses are performable from all the biological samples containing DNA. Thus, DNA extraction is the first and probably one of the most crucial steps of any genetic test. This chapter provides protocols for DNA extraction from both fresh blood and the big variety of biological samples that can be studied in the Forensic Laboratory. First, the general precautions which have to be adopted to prevent cross-contamination of the samples are underlined. All the extraction protocols (based upon Proteinase K/SDS lysis followed by ethanol precipitation) are carefully described while crucial steps are pointed out by footnotes. In addition, since PCR failure is common when handling aged/forensic specimens, strategies to remove substances acting as PCR inhibitors are described as well. At least, a simple and inexpensive PCR-based method allowing discriminating between removable and un-removable PCR inhibition is shown

Acquisition of different carbapenem resistance mechanisms by an epidemic clonal lineage of Pseudomonas aeruginosa

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Molecular evolution of metallo-beta-lactamase-producing Pseudomonas aeruginosa in a nosocomial setting of high-level endemicity

An outbreak of multidrug-resistant P. aeruginosa strains producing VIM-type metallo-beta-lacta-mases (MBLs) has occurred in an Italian hospital since 2000. In this work we characterized 128 carbapenem-resistant isolates collected from that hospital from 2000 to 2002. Genotyping showed that most VIM-positive isolates belonged to two different clonal lineages, producing either a VIM-1- or a VIM-2-like MBL. The 86 clonally related isolates carrying a blaVIM-1-like gene on an In70-like integron were clearly related to a VIM-1-positive P. aeruginosa clone circulating in various Italian hospitals since the late 1990s. In the VIM-2-like positive clone, the MBL gene was carried by an unusual class 1 integron, named In71, lacking the 3\u2019 conserved sequence region typical of sul1-associated integrons. A retrospective investigation did not reveal the presence of strains related to any of the VIM-producing isolates earlier than 1997