Epidemiological aspects of healthcare-associated infections and microbial genomics.

Hospital-acquired infections (HAIs) are a cause of continuously increasing morbidity and mortality. Most of these infections are caused by a limited set of bacterial species, which share the capability to efficiently spread from patient to patient and to easily acquire antibiotic resistance determinants. This renders correct and rapid species identification and antibiotic susceptibility testing (AST) important and underscores the relevance of bacterial epidemiological typing. The latter is needed for the sensitive detection and exact tracing of nosocomial spread of these potentially multidrug-resistant microorganisms (MDRO). Many microbial typing technologies have been developed and put to some level of executive practice, but it seems that the continued evolution in methodology has currently reached an apex: there is likely to be scientific and practical consensus on the ultimate typing potential of bacterial whole-genome sequencing (WGS). The possibility to perform pan-genomic nucleotide-to-nucleotide comparisons between strains belonging to a single species and to detect even minute changes in nucleotide order will identify closely related organisms, while upon accumulation of such mutations, independent descend can be assumed. Calibration of difference levels [i.e. number of single nucleotide polymorphisms (SNPs)] into categories of inter-strain relatedness needs to be performed in order to generate robust, portable typing schemes. Here, we will briefly discuss the state of affairs regarding bacterial epidemiology based upon WGS, its relatedness with the nomenclature of former typing approaches and the continuing need for a global typing language.

Amplicon-based next-generation sequencing: an effective approach for the molecular diagnosis of epidermolysis bullosa.

BACKGROUND: Epidermolysis bullosa (EB) is caused by mutations in genes that encode proteins belonging to the epidermal-dermal junction assembly. Due to the extreme clinical/genetic heterogeneity of the disease, the current methods available for diagnosing EB involve immunohistochemistry of biopsy samples and transmission electron microscopy followed by single-candidate gene Sanger sequencing (SS), which are labour-intensive and expensive clinical pathways. OBJECTIVES: According to the recently published recommendations for the diagnosis and treatment of EB, the assessment of the mutational landscape is now a fundamental step for developing a comprehensive diagnostic path. We aimed to develop a customized, cost-effective amplicon panel for the complete and accurate sequencing of all the pathogenic genes already identified in EB, and to minimize the processing time required for the execution of the test and to refine the analysis pipeline to achieve cost-effective results from the perspective of a routine laboratory set-up. Next-generation sequencing (NGS) via the parallel ultra-deep sequencing of many genes represents a proper method for reducing the processing time and costs of EB diagnostics. MATERIALS AND METHODS: We developed an EB disease-comprehensive AmpliSeq panel to accomplish the NGS on an Ion Torrent Personal Genome Machine platform. The panel was performed on 10 patients with known genetic diagnoses and was then employed in eight family trios with unknown molecular footprints. RESULTS: The panel was successful in finding the causative mutations in all 10 patients with known mutations, fully confirming the SS data and providing proof of concept of the sensitivity, specificity and accuracy of this procedure. In addition to being consistent with the clinical diagnosis, it was also effective in the trios, identifying all of the variants, including ones that the SS missed or de novo mutations. CONCLUSIONS: The NGS and AmpliSeq were shown to be an effective approach for the diagnosis of EB, resulting in a cost- and time-effective 72-h procedure.

Clinical epidemiology of hand eczema in patients accessing dermatological reference centres: results from Italy.

BACKGROUND: Data on the epidemiological impact and clinical characteristics of chronic hand eczema in Southern Europe are lacking. OBJECTIVES: To estimate the prevalence of chronic hand eczema in its different stages of severity and refractoriness to standard therapy in patients accessing Italian dermatological reference centres, and to evaluate sociodemographic and clinical factors associated with each stage. METHODS: A cross-sectional multicentre study was conducted. Adult patients with hand eczema, consecutively accessing 14 centres over a 6-month period, were enrolled. Patients were classified according to disease duration, severity and response to standard therapy with potent topical corticosteroids. Logistical regression was performed to investigate the relationship between sociodemographic and clinical data with different stages of eczema. RESULTS: The total number of participants was 981. Hand eczema was chronic in 83·5% of patients; 21·3% had severe eczema, with 62·0% of these patients refractory to standard therapy. Food processing and related work, the health professions, craft and related trade works (building, plumbing, electrical), hairdressing/beauty and handicraft work were most frequently associated with chronic hand eczema. Severe chronic hand eczema was more likely to be seen in men, older patients and those with less education. Severe and refractory hand eczema was also more likely among the unemployed and patients with allergic rhinitis and/or atopic dermatitis. CONCLUSIONS: Chronic hand eczema is frequent among patients with hand eczema accessing dermatology centres. Many patients were severe and refractory to standard therapy. The appropriate identification of hand eczema is the first step in implementing effective and efficient treatments.

Skin prick test: the only predictive tool of anaphylaxis? A case report.

Currently, in the literature there is a lack of definite predictive values parameters to identify patients with the risk to develop anaphylaxis. The controlled oral food challenge remains the gold standard for food allergy diagnosis. We report a case of a girl allergic to cow's milk with low levels of specific IgE and large skin prick test wheal sizes for cow's milk. In some cases the high diameter of skin prick test wheal may be more reliable than specific IgE levels in predicting an anaphylactic reaction.

Latex-fruit syndrome in Italian children and adolescents with natural rubber latex allergy.

Approximately 30-50 percent of individuals with natural rubber latex (NRL) allergy show an associated hypersensitivity to particular plant-derived foods, which has been defined "latex-fruit syndrome" (LFS). In our population of 22 patients with IgE-mediated NRL allergy we found a relevant prevalence (36 percent) of LFS, which resulted significantly higher in the group of patients with more severe clinical manifestations of NRL allergy than in patients with contact symptoms due to NRL (78 percent vs 8 percent; less than 0.005).

Oligonucleotide optical switches for intracellular sensing.

Fluorescence imaging coupled with nanotechnology is making possible the development of powerful tools in the biological field for applications such as cellular imaging and intracellular messenger RNA monitoring and detection. The delivery of fluorescent probes into cells and tissues is currently receiving growing interest because such molecules, often coupled to nanodimensional materials, can conveniently allow the preparation of small tools to spy on cellular mechanisms with high specificity and sensitivity. The purpose of this review is to provide an exhaustive overview of current research in oligonucleotide optical switches for intracellular sensing with a focus on the engineering methods adopted for these oligonucleotides and the more recent and fascinating techniques for their internalization into living cells. Oligonucleotide optical switches can be defined as specifically designed short nucleic acid molecules capable of turning on or modifying their light emission on molecular interaction with well-defined molecular targets. Molecular beacons, aptamer beacons, hybrid molecular probes, and simpler linear oligonucleotide switches are the most promising optical nanosensors proposed in recent years. The intracellular targets which have been considered for sensing are a plethora of messenger-RNA-expressing cellular proteins and enzymes, or, directly, proteins or small molecules in the case of sensing through aptamer-based switches. Engineering methods, including modification of the oligonucleotide itself with locked nucleic acids, peptide nucleic acids, or L-DNA nucleotides, have been proposed to enhance the stability of nucleases and to prevent false-negative and high background optical signals. Conventional delivery techniques are treated here together with more innovative methods based on the coupling of the switches with nano-objects.

Metabolic abnormalities associated with initiation of systemic treatment for psoriasis: evidence from the Italian Psocare Registry.

OBJECTIVE: To evaluate variations in laboratory parameters and diagnoses of selected clinical conditions up to 16 weeks after starting a new systemic psoriasis treatment for Psocare Registry enrollees. DESIGN: Prospective cohort study. SETTING: Italian public referral centres for psoriasis treatment. PATIENTS: First-time recipients (n = 10,539) of continuous systemic psoriasis treatment for at least 16 weeks. MAIN OUTCOME MEASURE: Mean variations in (weeks 8 and 16) and proportions of patients reaching a clinically meaningful increase in serum levels (week 16) of total and low-density lipoprotein cholesterol, triglycerides, aspartate amino transferase, alanine amino transferase and creatinine, as well as week-16 cumulative incidences of new diagnoses of diabetes mellitus and arterial hypertension. RESULTS: Mean cholesterol and triglyceride levels significantly increased in patients treated with acitretin or cyclosporine. Mean triglyceride levels also increased in efalizumab- and etanercept-treated patients. Mean transaminase values increased in methotrexate-treated patients, and mean aspartate amino transferase levels increased in infliximab-treated patients. The average serum creatinine value increased in cyclosporine-treated patients. Acitretin and cyclosporine were associated with risk of hypercholesterolaemia (odds ratios 1.51 and 1.34) and acitretin with risk of hypertriglyceridaemia (odds ratio 1.43). Methotrexate and infliximab were associated with risk of more than doubling the upper normal aspartate amino transferase (odds ratios 2.06 and 1.87) and alanine amino transferase (odds ratios 2.38 and 1.74) values. The relative risk of developing arterial hypertension and diabetes was increased for patients receiving cyclosporine (odds ratios 3.31 and 2.88). CONCLUSION: Systemic treatments for psoriasis resulted in heterogeneous effects on the parameters analysed.

Prolactinoma: a condition associated with hypoadiponectinemia.

Prolactinomas are prolactin-secreting neoplasias accounting for 40% of the pituitary adenomas. Much is known about the effects of prolactinomas on the reproductive system, but few data are yet available regarding their induced changes on metabolism. This study was aimed at evaluating patients with prolactinomas for insulin resistance and adiponectinemia. Forty patients with prolactinoma were allocated to 2 different groups according to disease control: 20 with uncontrolled disease (UPRL) and 20 with controlled disease in the last 6 months (CPRL). Forty healthy individuals (CG) matched for age, sex, and body mass index were taken as controls. Patients with prolactinoma were compared both as a one group and according to disease control with CG. All subjects were evaluated for waist/hip ratio (WHR), blood pressure, lipid profile, fasting glucose, homeostasis assessment model of insulin resistance (HOMAIR), and adiponectin. Patients with prolactinomas (UPRL+CPRL) showed higher insulin (p<0.05) and HOMAIR (p<0.05), alongside with lower adiponectin levels (p<0.01) than matched controls. When UPRL was compared to CPRL and CG, UPRL was disclosed as a subgroup of significant altered metabolic profile as related to WHR (p<0.01 for comparisons), high-density lipoprotein cholesterol (p<0.05 for comparisons), triglycerides (p<0.05 for comparisons), HOMAIR (p<0.05 and p<0.01, respectively), and adiponectin (p<0.01 for comparisons). All these metabolic abnormalities, except hypoadiponectinemia (p<0.01), were not observed in CPRL. These data suggest that prolactinomas are associated with hypoadiponectinemia, which is further exacerbated in uncontrolled patients when insulin resistance is also prominent.

Optical fibre gratings as tools for chemical and biochemical sensing.

Optical fibre gratings have recently been suggested as optical platforms for chemical and biochemical sensing. On the basis of the measurement of refractive index changes induced by a chemical and biochemical interaction in the transmission spectrum along the fibres, they are proposed as a possible alternative to the other label-free optical approaches, such as surface plasmon resonance and optical resonators. The combination of the use of optical fibres with the fact that the signal modulation is spectrally encoded offers multiplexing and remote measurement capabilities which the other technology platforms are not able to or can hardly offer. The fundamentals of the different types of optical fibre gratings are described and the performances of the chemical and biochemical sensors based on this approach are reviewed. Advantages and limitations of optical fibre gratings are considered, with a look at new perspectives for their utilization in the field.

Evolution of hypogammaglobulinemia in premature and full-term infants.

There are few data in the literature reporting the evolution of hypogammaglobulinemia in premature and full-term infants during the first years of life. The aim of this study was to assess the clinical and immunological evolution of premature and full-term infants with hypogammaglobulinemia. We included 24 children (11 premature and 13 full-term infants), aged 0-36 months, with hypogammaglobulinemia. Fifteen (62.5%) children had an isolated reduction in IgG, 7 (29.2%) had a decrease in both IgG and IgA and 2 (8.3%) a reduction in IgG and IgM. Normalization of IgG serum levels occurred in the premature infants at a mean age of 7.2 months. Full-term infants were divided into 3 groups based on age at normalization of IgG serum level: A) hypogammaglobulinemia with normalization within 12 months of life; B) with normalization within 36 months of life; C) normalization after 36 months. All the premature infants with hypogammaglobulinemia recovered, even though in the lower limits for age in the first years, while transient hypogammaglobulinemia observed in full-term infants has a different age of recovery.

Psoriasis relapse evaluation with week-end cyclosporine A treatment: results of a randomized, double-blind, multicenter study.

Cyclosporine A (CsA) effectively controls psoriasis, however, its long-term continuous use is not recommended. This study aims to evaluate the efficacy and tolerability of week-end CsA microemulsion for the reduction of relapse rate in patients with chronic plaque psoriasis who had achieved clinical remission following continuous CsA therapy. The PREWENT (Psoriasis Relapse Evaluation with Week-End Neoral Treatment) study was a 24 week, randomized, double-blind, multicenter study, carried out in 22 Italian hospital or University Dermatology units. CsA was discontinued for 8 days previous to the patients being randomized to oral CsA 5 mg/kg/day or placebo for two consecutive days/week, for a total period of 24 weeks. The primary endpoint was clinical success rate at week 24, defined as the proportion of patients with no clinical worsening (no relapse or a Psoriasis Area and Severity Index (PASI) < 75 percent of pre-treatment PASI). A total of 162 patients were randomized to CsA and 81 to placebo. Clinical success rates at 24 weeks were 66.9 and 53.2 percent with CsA and placebo, respectively (p = 0.072). Time to first relapse was significantly prolonged with CsA versus placebo (p = 0.023), and PASI was significantly lower from weeks 4 to 16 in CsA recipients. In patients with moderate-severe psoriasis, the clinical success rate was significantly increased with CsA compared to placebo (69.9% vs 46.3%; p = 0.011), and significantly lower increases in PASI were observed from week 4 to week 24 (p < 0.05 vs placebo). CsA was well tolerated, with no differences in mean blood creatinine or blood pressure between CsA and placebo recipients. However, the high withdrawal rate (22.2% of randomized patients), which was not related to side effects, may have led to an overestimation of efficacy, but the study had a good statistical power (88% greater than that observed in similar studies, i.e. 80%). Week-end CsA administration was shown to prolong safely and effectively the time to first relapse in psoriasis patients.

Evolution of and risk factors for psychological distress in patients with psoriasis: the PSYCHAE study.

Psychological distress (PD) is common in patients with psoriasis but little is known about its evolution. The aim of this study is to assess the evolution of PD in psoriasis. For this purpose, 1,505 psoriatic patients, who had been previously enrolled in the PSYCHAE study, an observational multicenter Italian study, were re-evaluated after 6 and 12 months. Minor and major PD were assessed using the General Health Questionnaire (GHQ) and Brief Symptoms Inventory (BSI) questionnaires and coping using Brief COPE questionnaire. Minor PD was present in 46 percent of patients but halved during the study. Female gender, surface area, topical steroids, methotrexate, self-distraction, venting and behavioral disengagement were risk factors for minor PD; cyclosporine and humor were protective. Major PD was present in 11 percent of patients and remained stable. Female gender, venting, religion, behavioral disengagement and emotional support were risk factors for major PD; instrumental support was protective. In conclusion, the results obtained suggest that major PD remained stable after 12 months and that coping was a predictor of its evolution.

Allergy is not the main trigger of urticaria in children referred to the emergency room.

BACKGROUND: Urticaria is the disease that has the highest impact on quality of life and requires the most visits to the emergency room. OBJECTIVE: To investigate the clinical presentation of acute urticaria in children referred to the paediatric emergency room of our hospital and to define possible related aetiologies. METHODS: We included 814 children consecutively referred to the emergency room between January 2006 and December 2007 with a diagnosis of acute urticaria, isolated or associated with other clinical symptoms. RESULTS: Only 2.0% of the cases studied were associated with severe clinical pictures. In 437 cases (53.7%), the cause of urticaria was not determined. The infections of the respiratory tract were the most frequently suspected aetiological factor. The diagnosis of allergic urticaria is more defined, but belongs to a minority group (10.8%). The first level treatment includes the use of non-sedating oral H1-antihistamine. CONCLUSION: The children with urticaria are frequently referred to the paediatric emergency room, but only in a few cases were associated with severe clinical manifestations or allergy. The evidence of an inverse relationship between the number of accesses and the patients' age may be explained by the higher prevalence of this disease in early childhood and possibly also by a higher concern of the parents of the younger patients.

An Italian shared dermatological and rheumatological proposal for the use of biological agents in psoriatic disease.

BACKGROUND: As psoriatic disease (PD) is a condition characterized by the combination of inflammatory skin (psoriasis) and osteo-articular manifestations (psoriatic arthritis), its treatment should cover both its clinical components. OBJECTIVE: The objective of this study was to propose a flexible framework for the use of biological agents in PD. METHODS: The proposal was drawn up by a group of dermatologists and rheumatologist expert in PD and was based on existing evidence and personal opinion. RESULTS: The three TNF-alpha inhibitors (adalimumab, etanercept, infliximab) are effective in all of the psoriatic manifestations and should be used in the case of moderate/severe disease refractory to systemic treatment with non-biological drugs. We propose the following definitions of moderate/severe disease: PASI > 10 or BSA > 10 or DLQI > 10 for plaque-psoriasis; BSA > or = 10 or DLQI > or = 10 for the other psoriatic skin lesions; DLQI > or = 10 or meaningful values of the NAPSI or mNAPSI for psoriatic nail involvement; > or = 1 inflamed joint + patient global VAS(3)4 + physician's judgement or arthritic joint deformities or radiographic joint damage plus > or = 5 inflamed small joints or > or = 1 large joints for peripheral joint involvement; > or = 1 digit with dactylitis and > or = 1 enthesitic sites + patient global VAS(3)4 + physician's judgement for dactylitis and enthesitis. BASDAI > or = 4 + physician's judgement for spondylitis; recurrent flares or risk of developing irreversible damage for uveitis. Other assessment instruments can be used if the physician is more familiar with them and if they have been validated. CONCLUSION We provide a shared dermatological and rheumatological proposal for the use of biological agents in PD.

High-Q polymer-coated microspheres for immunosensing applications.

Homogeneous polymeric thin layers have been used as functionalizing agents on silica microspherical resonators in view of the implementation of an immunosensor. We have characterized the microspheres functionalized with poly-L-lactic acid and Eudragit L100, as an alternative to the commonly used 3-Aminopropyltrimethoxysilane. It is shown that polymeric functionalization does not affect the high quality factor (Q greater than 10(7)) of the silica microspheres, and that the Q factor is about 3 x 10(5) after chemical activation and covalent binding of immunogammaglobulin (IgG). This functionalizing process of the microresonator constitutes a promising step towards the achievement of an ultra sensitive immunosensor.

Transforming growth factor-beta 1 modulates beta 1 and beta 5 integrin receptors and induces the de novo expression of the alpha v beta 6 heterodimer in normal human keratinocytes: implications for wound healing.

The molecular mechanism underlying the promotion of wound healing by TGF-beta 1 is incompletely understood. We report that TGF-beta 1 regulates the regenerative/migratory phenotype of normal human keratinocytes by modulating their integrin receptor repertoire. In growing keratinocyte colonies but not in fully stratified cultured epidermis, TGF-beta 1: (a) strongly upregulates the expression of the fibronectin receptor alpha 5 beta 1, the vitronectin receptor alpha v beta 5, and the collagen receptor alpha 2 beta 1 by differentially modulating the synthesis of their alpha and beta subunits; (b) downregulates the multifunctional alpha 3 beta 1 heterodimer; (c) induces the de novo expression and surface exposure of the alpha v beta 6 fibronectin receptor; (d) stimulates keratinocyte migration toward fibronectin and vitronectin; (e) induces a marked perturbation of the general mechanism of polarized domain sorting of both beta 1 and beta 4 dimers; and (f) causes a pericellular redistribution of alpha v beta 5. These data suggest that alpha 5 beta 1, alpha v beta 6, and alpha v beta 5, not routinely used by keratinocytes resting on an intact basement membrane, act as "emergency" receptors, and uncover at least one of the molecular mechanisms responsible for the peculiar integrin expression in healing human wounds. Indeed, TGF-beta 1 reproduces the integrin expression pattern of keratinocytes located at the injury site, particularly of cells in the migrating epithelial tongue at the leading edge of the wound. Since these keratinocytes are inhibited in their proliferative capacity, these data might account for the apparent paradox of a TGF-beta 1-dependent stimulation of epidermal wound healing associated with a growth inhibitory effect on epithelial cells.

A new procalcitonin optical immunosensor for POCT applications.

A new immunosensor for the determination of procalcitonin was developed. A sandwich assay format was implemented on a polymethylmetacrylate optical biochip, opportunely shaped in order to obtain several flow channels and potentially suitable for point of care testing applications. The sandwich format makes use of two new rat monoclonal antibodies. The capture antibody was covalently immobilised on the surface of the plastic chip, and the detection antibody was labelled with DY647 dye. Different combinations of capture and detection antibodies were investigated, and particular attention was devoted in order to avoid the non-specific adsorption. A limit of detection of 0.088 mg L(-1) was achieved within the working range of 0.28-50 mg L(-1) in buffer samples. The assay was also implemented in human serum, and 0.2 and 0.7-25 mg L(-1) were the attained limit of detection and working range, respectively.

Italian guidelines and therapeutic algorithm for actinic keratoses.

The prevalence of actinic keratosis (AK) continues to rise among white people throughout the world and it is necessary to increase the level of attention paid to it from a diagnostic and a preventive point of view. Today, AK must be considered an in situ squamous cell carcinoma and as such, must be managed using one of the available approved therapeutic alternatives. However, when multiple AKs develop on severely photodamaged skin, the treatment of the lesion together with that of the field of cancerization is part of an optimal strategy that aims not only to solve alterations clinically evident but also those in the surrounding skin field cancerization, that most likely hosts genetic alterations and is the site of initial gradual replacement of normal cells with tumoral cells. This paper reports the most recent evidences from a careful review of the literature's key articles of the treatment of AKs and suggests guidelines for the clinicians. The guidelines indicated by the authors have also been based on practical evaluations and their own clinical experience. The present conclusions may be modified by new findings in the field of oncologic research.