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To highlight the impact of peak height analysis on the inference to be drawn from typical, two persons mixtures (2PMs), a number of real high-quality – either "balanced" or "unbalanced" – 2PMs were subjected to a classical statistical analysis based on the likelihood of a known contributor to be therein. We calculated a LR value based on the genetic markers belonging to a real contributor (S1) and to a number of false but compatible suspects using a discrete/semi-continuous method (as implemented in a shareware of widespread adoption[LR-mix Studio]) and a quantitative approach of our own conception (built into a Microsoft Excel spreadsheet). Quantitative information conferred by the height of the peaks was irrelevant in the case of "balanced" mixtures but played a key role in eliciting correct inference from "unbalanced" 2PMs.

Since 1992, the Spanish and Portuguese-Speaking Working Group of the ISFG (GHEP-ISFG) has been organizing annual Intercomparison Exercises (IEs) coordinated by the Quality Service at the National Institute of Toxicology and Forensic Sciences (INTCF) from Madrid, aiming to provide proficiency tests for forensic DNA laboratories. Each annual exercise comprises a Basic (recently accredited under ISO/IEC 17043: 2010) and an Advanced Level, both including a kinship and a forensic module. Here, we show the results for both autosomal and sex-chromosomal STRs, and for mitochondrial DNA (mtDNA) in two samples included in the forensic modules, namely a mixture 2:1 (v/v) saliva/blood (M4) and a mixture 4:1 (v/v) saliva/semen (M8) out of the five items provided in the 2014 GHEP-ISFG IE. Discrepancies, other than typos or nomenclature errors (over the total allele calls), represented 6.5% (M4) and 4.7% (M8) for autosomal STRs, 15.4% (M4) and 7.8% (M8) for X-STRs, and 1.2% (M4) and 0.0% (M8) for Y-STRs. Drop-out and drop-in alleles were the main cause of errors, with laboratories using different criteria regarding inclusion of minor peaks and stutter bands. Commonly used commercial kits yielded different results for a micro-variant detected at locus D12S391. In addition, the analysis of electropherograms revealed that the proportions of the contributors detected in the mixtures varied among the participants. In regards to mtDNA analysis, besides important discrepancies in reporting heteroplasmies, there was no agreement for the results of sample M4. Thus, while some laboratories documented a single control region haplotype, a few reported unexpected profiles (suggesting contamination problems). For M8, most laboratories detected only the haplotype corresponding to the saliva. Although the GHEP-ISFG has already a large experience in IEs, the present multi-centric study revealed challenges that still exist related to DNA mixtures interpretation. Overall, the results emphasize the need for further research and training actions in order to improve the analysis of mixtures among the forensic practitioners.

Objectives To analyze Staphylococcus epidermidis strains, previously tested for their virulence in a mouse model of subcutaneous infection, for various phenotypic traits (biofilm density, extracellular polysaccharide, slime-associated antigen (SAA)) and for the presence of the ica gene cluster, to determine which of these phenotypic and genotypic methods best correlates with virulence in the mouse model. Methods The quantitative biofilm assay was performed on 10 strains of S. epidermidis , comprising (1) RP62A (ATCC 35984), (2) the strongest and weakest biofilm producers in our collection, (3) a pair of phenotypic variants, and (4) a strain whose biofilm density was enhanced in iron-limited media. Biofilm density was measured after growth at 37 °C and at ambient temperature, in trypticase soy broth (TSB) with and without glucose supplementation and using both chemical and heat fixation. Strains were assayed for SAA using a double immunodiffusion method. Extracellular polysaccharide was detected by transmission electron microscopy (TEM). A 546-base-pair segment of the ica gene cluster was amplified by PCR. Results Biofilm formation in TSB, glucose-enriched TSB, extracellular polysaccharide (observed by TEM), expression of SAA and presence of the ica gene predicted virulence of nine, nine, nine, eight and eight of 10 strains, respectively. The phenotypic expression of biofilm and related properties was medium and temperature dependent. We encountered one ica -positive strain that failed to express biofilm in standard TSB at 37 °C, but was virulent in a mouse model, and another strain that lacked ica , produced biofilm and was virulent in the model. Conclusions Mouse virulence in our model can be predicted by any of the phenotypic or genotypic methods examined for ≥80% of strains. Medium and incubation conditions affect the expression of phenotypic markers by some strains. For the remaining strains, possible reasons for inconsistencies between the presence of the ica gene, phenotypic markers and mouse virulence include (1) dependence of biofilm on genes other than ica , (2) sequence differences in ica , (3) dependence of biofilm expression in vivo on strain characteristics and media used to prepare inocula for in vivo studies.

The development of new antimicrobial therapeutic tools addresses the emergence of multidrug-resistant micro-organisms or clones and the need for more effective antimicrobial strategies. Overcoming the hurdles in providing early diagnosis and intervention on hard-to-reach and/or resting bacteria (i.e. biofilm-embedded cells) represents a challenging task. In this review, we identify a set of organic, inorganic, and hybrid materials that might be used for prevention and control of healthcare-associated infections. We report the current knowledge on nano- and microparticle-based antimicrobial agents and describe the possible mode of their action.

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.

The past 2 decades have brought worrying increases in severe Streptococcus pyogenes diseases globally. To investigate and compare the epidemiological patterns of these diseases within Europe, data were collected through a European Union FP-5-funded program (Strep-EURO). Prospective population-based surveillance of severe S. pyogenes infection diagnosed during 2003 and 2004 was undertaken in 11 countries across Europe (Cyprus, the Czech Republic, Denmark, Finland, France, Germany, Greece, Italy, Romania, Sweden, and the United Kingdom) using a standardized case definition. A total of 5,522 cases were identified across the 11 countries during this period. Rates of reported infection varied, reaching 3/100,000 population in the northern European countries. Seasonal patterns of infection showed remarkable congruence between countries. The risk of infection was highest among the elderly, and rates were higher in males than in females in most countries. Skin lesions/wounds were the most common predisposing factor, reported in 25% of cases; 21% had no predisposing factors reported. Skin and soft tissue were the most common foci of infection, with 32% of patients having cellulitis and 8% necrotizing fasciitis. The overall 7-day case fatality rate was 19%; it was 44% among patients who developed streptococcal toxic shock syndrome. The findings from Strep-EURO confirm a high incidence of severe S. pyogenes disease in Europe. Furthermore, these results have identified targets for public health intervention, as well as raising awareness of severe S. pyogenes disease across Europe. Copyright © 2008, American Society for Microbiology. All Rights Reserved.

The effect of a nonsteroidal antiinflammatory drug commercially available in eye drop form (sodium diclofenac) was assayed for its ability to affect biofilms formed by clinical Staphylococcus epidermidis isolates. Biofilms produced by one strain positive for a slime-associated antigen, suggested to be expressed by more virulent strains, was not affected by sodium diclofenac treatment. On the other hand, biofilm produced by the slime-positive, antigen-negative strain showed dramatic alterations already after short treatments with sodium diclofenac as reported for salicylate and other nonsteroidal drugs. Such results suggest further investigation of the possible use of sodium diclofenac drops in the treatment of ophthalmic infections in soft contact lens wearers.

A case of prurigo nodularis (Hyde's prurigo) preceding by 15 years the diagnosis of celiac disease is presented. In particular, the association between the clinical course of prurigo nodularis and the compliance with gluten-free diet is reported and discussed.

Publisher Summary This chapter presents the basic approaches to counting microorganisms on a plastic surface, which is the material of choice for the fabrication of most medical appliances. Much of the discussion presented in the chapter is also relevant to the study of the colonization of non-plastic materials by environmental organisms and non-plastic medical appliances by pathogenic bacteria. Like the colonization of tissues, the colonization of medical devices is a multiphasic process. Colonization begins with either a directed or a random encounter of the microbe with the substratum followed by the rapid binding of the microbe to the surface. Microbial proliferation, the penultimate phase of colonization, is subject to the concentration of nutrients in the bathing fluid and the presence of other microorganisms on the surface. When colonization takes place in vivo, microbial multiplication may also be subject to the humoral and cellular immune responses of the host as well as the administration of antimicrobial agents. Colonization is concluded by an often-overlooked stage in which there is release of free-floating (planktonic) organisms from the sessile colony and dispersal of the planktonic forms to new colonization sites. Release occurs when daughter cells stop expressing adhesive structures. This may be accomplished either by a cellular response to a change in environmental conditions (phenotypic modulation) or by a (usually reversible) spontaneous change in the genetic mechanisms that govern the expression of microbial adhesive structures (phase variation).

Infections is one of the most common cause of catheter failure as well as the most difficult to manage, most often requiring catheter removal. Staphylococcus is the etiologic agent of such infections more frequently isolated, particularly Staphylococcus epidermidis. Several factors have been suggested to be involved in bacteria-biomaterial interactions such as catheter surface morphology, molecular biofilm and bacterial virulence features. Different strategies have been tried to avoid the development to catheter-associated infections: among them adsorption of antibiotic molecules to the catheter surface might represent a successful tool to improve catheter implant life.

Gonococcal adherence was studied in vitro using buccal epithelial cells (BEC). In smears stained with the Gram method, a progressive decrease in gonococcal adherence to the BEC after some culture passages was observed. There was a parallel decrease to almost total disappearance in the number of fimbriated bacteria. An electron microscopy study showed that adherence to the epithelial cells was mediated by fimbriae and, in part, by a polysaccharide component of the bacterial cell wall which seems to guarantee persistent adherence ability, even after the loss of fimbriae.

Fourteen strains of Clostridium difficile, previously characterized by SDS-PAGE, were examined for the presence of surface structures. None of the strains were fimbriated but, when grown in the presence of glucose, all exhibited a thin ruthenium red-positive layer. Two strains, belonging to different electrophoretic groups, were also observed after treatment with homologous and heterologous antisera. The strain belonging to the electrophoretic group 2, usually associated with the disease, showed a 30-80nm thick capsule in ultrathin sections. The strains belonging to group 5, mainly observed in isolates from healthy children, exhibited a thinner polysaccharide layer (10-20 nm). No stabilization was observed when these strains were treated with heterologous antisera. This capsule-like material did not seem to be associated with adhesive properties of C. difficile strains.

Five strains of Clostridium spiroforme were examined for their surface properties. All strains were able to agglutinate human erythrocytes. Electron microscopy showed a ruthenium red-positive capsule mediating the attachment of bacteria to erythrocytes. Two strains, showing the lowest degree of haemagglutination, exhibited an additional external layer of filamentous structures, possibly interfering with the agglutinating activity. In spite of their agglutinating ability, the C. spiroforme strains did not show surface hydrophobicity, thus suggesting the possible existence of a new type of clostridial adhesin.

The penetration process of an enteroinvasive strain of Escherichia coli 0124 into HEp-2 cells has been studied by electron microscopy. Bacteria penetrated into cultured cells by a phagocytosis-like mechanism, and multiplied within the vacuoles as well as in the cytoplasmic matrix. On the whole, the infection cycle of EIEC appeared to be similar to that described for Shigella. The stain examined has also been found to elaborate a 40 nm-thick glyocalix, which is probably involved in the recognition of host surface receptors and therefore in the attachment of microorganisms to the cell membrane.