A novel versatile precursor suitable for 18 F-radiolabeling via "click chemistry".

As an effort to improve 18 F-radiolabeling of biomolecules in method robustness and versatility, we report the synthesis and radiolabeling of a new azido precursor potentially useful for the so-called "click reaction," in particular the ligand-free version of the copper(I)-catalyzed alkyne-azide cycloaddition. The new azido precursor may help to overcome problems sometimes exhibited by most of the currently used analogues, as it is safe to handle and it displays long-term chemical stability, thus facilitating the development of new radiolabeling procedures. Moreover, the formed 18 F-labeled 1,2,3-triazole is potentially metabolically stable and could enhance the in vivo circulation time. The above azido precursor was successfully radiolabeled with 18 F, with 51% radiochemical yield (nondecay-corrected). As a proof of concept, the 18 F-labeled azide was then tested with a suitable alkyne functionalized aminoacid (l-propargylglycine), showing 94% of conversion, and a final radiochemical yield of 27% (>99% radiochemical purity), nondecay-corrected, with a total preparation time of 104 minutes.

NaCl concentration in the medium modulates the secretion of active EmpA protease in Vibrio anguillarum at post-transcriptional level.

AIMS: This study focused on the influence of different amounts of NaCl in the medium in Vibrio anguillarum EmpA protease production at both the transcriptional and post-transcriptional levels. METHODS AND RESULTS: Vibrio anguillarum 975/I was cultivated in cM9 medium with varying concentrations of NaCl: 0·5, 1·5, 3·0%. EmpA protease was monitored in the supernatants by the skim milk test, azocasein assay and Western blot analysis. The empA gene expression was measured by real-time PCR. A mutant strain 975/I defective for the empA gene confirmed the specificity of the response for EmpA protease. Active protease production was induced by 0·5 and 1·5% NaCl-amended media; however, the strain cultivated in 3·0% NaCl was unable to secrete EmpA protease. The quantitative expression of the empA gene was very similar in all tested conditions. CONCLUSIONS: The NaCl concentration in the medium modulates the secretion of active EmpA protease in V. anguillarum at a post-transcriptional level. SIGNIFICANCE AND IMPACT OF THE STUDY: EmpA protease is one of the most important virulence factors in V. anguillarum. We demonstrated the influence of osmotic changes in the regulation of EmpA protease in the V. anguillarum 975/I strain. This finding has an important impact on the evaluation of factors determining the onset of disease in fish.

Allergen sensitizations in southern Italy: a 5-year retrospective study in allergic respiratory patients.

The assessment of the distribution of allergen skin test sensitizations is highly recommended for the optimal management of allergic respiratory conditions. We aimed at evaluating the distribution of allergen sensitizations in individuals with asthma and/or rhinitis in the Southern region of ltaly, and at exploring whether changes in the frequency of allergen sensitizations occurred after a 5-year period. Demographic data and skin prick test sensitizations to allergens from asthmatics and/or rhinitis attending the Division of Respiratory Diseases, University of Palermo, Italy in 2005 (Phase 1) and in 2010 (Phase 2) were extrapolated and retained for analysis. A total of 2033 allergic respiratory patients were included (1002 in Phase 1 and 1031 in Phase 2). In both investigations, the most prevalent allergen sensitization was towards Parietaria; however, a significant reduction in the rate of prevalence after 5 years was recorded (from 60% to 48% of skin test positive patients, p < 0.0001). Up to one out of two subjects showed sensitization to dust mites in both Phases. Interestingly, Cypress pollen sensitization almost doubled from Phase 1 (17%) to Phase 2 (29%; p < 0.0001). Overall, the mean number of skin test sensitizations for each patient increased from 2.7 +/- 1.6 in Phase 1 to 3.1 + 1.8 in Phase 2 (p < 0.0001). The present findings confirm the prevalent role of Parietaria sensitization in the allergic population of the Mediterranean area of Southern Italy, and document the increase of Cypress sensitization. These observations could contribute to a proper management of chronic allergic respiratory conditions in this region.

Perspectives in linear accelerator for FLASH VHEE: Study of a compact C-band system.

PURPOSE: In order to translate the FLASH effect in clinical use and to treat deep tumors, Very High Electron Energy irradiations could represent a valid technique. Here, we address the main issues in the design of a VHEE FLASH machine. We present preliminary results for a compact C-band system aiming to reach a high accelerating gradient and high current necessary to deliver a Ultra High Dose Rate with a beam pulse duration of 3µs. METHODS: The proposed system is composed by low energy high current injector linac followed by a high acceleration gradient structure able to reach 60-160 MeV energy range. To obtain the maximum energy, an energy pulse compressor options is considered. CST code was used to define the specifications RF parameters of the linac. To optimize the accelerated current and therefore the delivered dose, beam dynamics simulations was performed using TSTEP and ASTRA codes. RESULTS: The VHEE parameters Linac suitable to satisfy FLASH criteria were simulated. Preliminary results allow to obtain a maximum energy of 160 MeV, with a peak current of 200 mA, which corresponds to a charge of 600 nC. CONCLUSIONS: A promising preliminary design of VHEE linac for FLASH RT has been performed. Supplementary studies are on going to complete the characterization of the machine and to manufacture and test the RF prototypes.

Sumo1-ylation of human spermatozoa and its relationship with semen quality.

Sumoylation is a post-translational modification involved in the regulation of several cell functions. Recent studies suggest its involvement in spermatogenesis, but occurrence and function of SUMO (small ubiquitin-like modifier) in mature spermatozoa remain unknown. We report the occurrence of several SUMO1-conjugated proteins, in a range of 20-85 kDa, in ejaculated spermatozoa. By cytofluorimetric analysis, we evaluated the percentage of SUMO1-positive spermatozoa in 58 subjects undergoing semen analysis in our laboratory and correlated the obtained values with semen parameters. We found that the percentage of SUMO1-positive spermatozoa was inversely correlated with total (r = -0.35, p < 0.01) and progressive motility (r = -0.29, p < 0.05). Such correlations become stricter when only asthenospermic subjects were included in the analysis (r = -0.58, p = 0.01 for progressive motility, n = 17) and were lost in non-asthenospermic subjects. By immunofluorescence and immunoconfocal fluorescence, we demonstrated that SUMO1 is mainly located in the nucleus and, occasionally, in the midpiece of spermatozoa. Immunoelectron microscopy as well as a long permeabilization protocol demonstrated a massive localization of SUMO-1 in the nucleus. By using a fluorescent probe to distinguish dead/live cells, we show that SUMO1 is mainly present in live spermatozoa. In conclusion, sumoylation of human spermatozoa may be involved in the regulation of motility.

Bioremediation of benzene, toluene, ethylbenzene, xylenes-contaminated soil: a biopile pilot experiment.

AIMS: In this study, we evaluated the removal efficiency of fuel hydrocarbons from a jet fuel contaminated area using bioaugmentation treatment in biopile. METHODS AND RESULTS: The hydrocarbon analysis of the sample revealed total hydrocarbons mainly constituted by benzene, toluene, ethylbenzene, xylenes (BTEX) and heavy aliphatic hydrocarbons. Enrichments of soil sample were performed with BTEX, pristane and fuel JP-5, respectively, selected hydrocarbon-degrading strains, namely Acinetobacter sp., Pseudomonas sp. and Rhodococcus sp. Three hundred litres of culture containing 10(8) cell ml(-1) of each strain and nutrients sprayed on the biopile allowed a removal of 90% of total hydrocarbons in 15 days. Bioremediation process was monitored by observation of the respiration rate and the bacterial abundance and GC-MS analysis. CONCLUSIONS: The efficiency of the treatment in the biopile was considerable. The assessment of microbial activity during the experiment is necessary for interventions targeted to improve environmental parameters such as humidity, temperature, pH and nutrients for optimization of the bioremediation process. SIGNIFICANCE AND IMPACT OF THE STUDY: A better knowledge of microbial successions at oil-polluted sites is essential for environmental bioremediation. Data obtained in biopile study improve our understanding of processes occurring during oil pollution.

Epidemiology, management, and outcome of carbapenem-resistant Klebsiella pneumoniae bloodstream infections in hospitals within the same endemic metropolitan area.

In the last decade, carbapenem-resistant Klebsiella pneumoniae (CR-Kp) has become endemic in several countries, including Italy. In the present study, we assessed the differences in epidemiology, management, and mortality of CR-Kp bloodstream infection (BSI) in the three main adult acute-care hospitals of the metropolitan area of Genoa, Italy. From January 2013 to December 2014, all patients with CR-Kp BSI were identified through the computerized microbiology laboratory databases of the three hospitals. The primary endpoints of the study were incidence and characteristics of CR-Kp BSI in hospitals within the same endemic metropolitan area. Secondary endpoints were characteristics of CR-Kp BSI in hospitals with and without internal infectious diseases consultants (IDCs) and 15-day mortality. During the study period, the incidence of healthcare-associated CR-Kp BSI in the entire study population was 1.35 episodes per 10,000 patient-days, with substantial differences between the three hospitals. Patients admitted to the two hospital with internal IDCs were more likely to receive post-susceptibility test combined therapy including carbapenems (77% vs. 26%, p<0.001), adequate post-susceptibility test therapies (86% vs. 52%, p<0.001), and post-susceptibility therapies prescribed by an infectious diseases specialist (84% vs. 14%, p<0.001). Overall, the crude 15-days mortality was 26%. In the final multivariable model, only septic shock at BSI presentation was unfavorably and independently associated with 15-days mortality (odds ratio [OR] 6.7, 95% confidence intervals [CI] 2.6-17.6, p<0.001), while a protective effect was observed for post-susceptibility test combined therapies including a carbapenem (OR 0.11, 95% CI 0.03-0.43, p=0.002). Mortality of CR-Kp remains high. Differences in the incidence of CR-Kp BSI were detected between acute-care centers within the same endemic metropolitan area. Efforts should be made to improve the collaboration and coordination between centers, to prevent further diffusion of CR-Kp.

Parotid gland involvement in Heerfordt syndrome: a case report.

Sarcoidosis is a multisystemic granulomatous disease characterized by the presence of noncaseating granulomas, the exact etiology of which is yet to be determined. Most of patients show granulomas located in the lungs or in the related lymph nodes. However, lesions can affect any organ. Noncaseating granulomas are not a pathognomonic sign of sarcoidosis, being observed also in other diseases, therefore the diagnosis is often of exclusion. We report a case of sarcoidosis with parotid gland involvement in the context of a Heerfordt syndrome, discussing about its clinical presentation, pathogenesis, pathology and differential diagnosis with other granulomatous diseases.

Ferrous iron- and ammonium-rich diffuse vents support habitat-specific communities in a shallow hydrothermal field off the Basiluzzo Islet (Aeolian Volcanic Archipelago).

Ammonium- and Fe(II)-rich fluid flows, known from deep-sea hydrothermal systems, have been extensively studied in the last decades and are considered as sites with high microbial diversity and activity. Their shallow-submarine counterparts, despite their easier accessibility, have so far been under-investigated, and as a consequence, much less is known about microbial communities inhabiting these ecosystems. A field of shallow expulsion of hydrothermal fluids has been discovered at depths of 170-400 meters off the base of the Basiluzzo Islet (Aeolian Volcanic Archipelago, Southern Tyrrhenian Sea). This area consists predominantly of both actively diffusing and inactive 1-3 meters-high structures in the form of vertical pinnacles, steeples and mounds covered by a thick orange to brown crust deposits hosting rich benthic fauna. Integrated morphological, mineralogical, and geochemical analyses revealed that, above all, these crusts are formed by ferrihydrite-type Fe3+ oxyhydroxides. Two cruises in 2013 allowed us to monitor and sampled this novel ecosystem, certainly interesting in terms of shallow-water iron-rich site. The main objective of this work was to characterize the composition of extant communities of iron microbial mats in relation to the environmental setting and the observed patterns of macrofaunal colonization. We demonstrated that iron-rich deposits contain complex and stratified microbial communities with a high proportion of prokaryotes akin to ammonium- and iron-oxidizing chemoautotrophs, belonging to Thaumarchaeota, Nitrospira, and Zetaproteobacteria. Colonizers of iron-rich mounds, while composed of the common macrobenthic grazers, predators, filter-feeders, and tube-dwellers with no representatives of vent endemic fauna, differed from the surrounding populations. Thus, it is very likely that reduced electron donors (Fe2+ and NH4+ ) are important energy sources in supporting primary production in microbial mats, which form a habitat-specific trophic base of the whole Basiluzzo hydrothermal ecosystem, including macrobenthic fauna.

Bioremediation technologies for polluted seawater sampled after an oil-spill in Taranto Gulf (Italy): A comparison of biostimulation, bioaugmentation and use of a washing agent in microcosm studies.

One of the main challenges of bioremediation is to define efficient protocols having a low environmental impact. We have investigated the effect of three treatments in oily-seawater after a real oil-spill occurred in the Gulf of Taranto (Italy). Biostimulation with inorganic nutrients allowed the biodegradation of the 73±2.4% of hydrocarbons, bioaugmentation with a selected hydrocarbonoclastic consortium consisting of Alcanivorax borkumensis, Alcanivorax dieselolei, Marinobacter hydrocarbonoclasticus, Cycloclasticus sp. 78-ME and Thalassolituus oleivorans degraded 79±3.2%, while the addition of nutrients and a washing agent has allowed the degradation of the 69±2.6%. On the other hand, microbial community was severely affected by the addition of the washing agent and the same product seemed to inhibit the growth of the majority of strains composing the selected consortium at the tested concentration. The use of dispersant should be accurately evaluated also considering its effect on the principal actors of biodegradation.

Biodegradation potentiality of psychrophilic bacterial strain Oleispira antarctica RB-8(T).

The present study is focused on assessing the growth and hydrocarbon-degrading capability of the psychrophilic strain Oleispira antarctica RB-8(T). This study considered six hydrocarbon mixtures that were tested for 22days at two different cultivation temperatures (4 and 15°C). During the incubation period, six sub-aliquots of each culture at different times were processed for total bacterial abundance and GC-FID (gas chromatography-flame ionization detection) hydrocarbon analysis. Results from DNA extraction and DAPI (4',6-diamidino-2-phenylindole) staining showed a linear increase during the first 18days of the experiment in almost all the substrates used; both techniques showed a good match, but the difference in values obtained was approximately one order of magnitude. GC-FID results revealed a substantial hydrocarbon degradation rate in almost all hydrocarbon sources and in particular at 15°C rather than 4°C (for commercial oil engine, oily waste, fuel jet, and crude oil). A more efficient degradation was observed in cultures grown with diesel and bilge water at 4°C.

A putative lichenysin A synthetase operon in Bacillus licheniformis: initial characterization.

Certain Bacillus licheniformis strains isolated from oil wells have been shown to produce a very effective biosurfactant, lichenysin A, which is structurally similar to another less active lipopeptide, surfactin. Surfactin, like many small peptides in prokaryotes and lower eukaryotes, is synthesized non-ribosomally by multi-enzyme peptide synthetase complex. Analysis of several peptide synthetases of bacterial and fungal origin has revealed a high degree of sequence conservation. Two 35-mer oligonucleotides derived from highly conserved motifs ('core I' and 'core II') of surfactin synthetase were used to identify the cloned putative operon of lichenysin A synthetase lchA from B. licheniformis BNP29, a strain not amenable to genetic manipulation in a BAC system (F-plasmid-based bacterial artificial chromosome) based on Escherichia coli and its single-copy plasmid F-factor. A 32.4 kb fragment containing lichenysin A biosynthesis locus was sequenced and analysed. The structural architecture of putative lichenysin A synthetase protein containing seven amino acid (aa) activation-thiolation, two epimerization and one thioesterase domains is discussed in terms of its similarity to surfactin and other peptide synthetases. The 100 aa peptide chain situated between the highly conserved signature sequences FDXX and NXYGPTE(IV)X within amino acid binding domains of peptide synthetases is proposed to be a minimal block dictating the substrate specificity of the enzymes. A new operon-type structure has been localized directly upstream from the lichenysin A synthetase genes which, on the basis of sequence determination, potentially encode a four-member ABC-type transport system involved in product secretion.

Reduced plasma ascorbic acid concentrations in nonsmokers regularly exposed to environmental tobacco smoke.

Oxidants in cigarette smoke accelerate metabolic turnover of ascorbic acid (AA), and thereby deplete body stores of this potent antioxidant and putative anticarcinogen in active smokers. We examined plasma AA concentrations and vitamin C intakes in nonsmokers regularly exposed to environmental tobacco smoke (ie, passive smokers; n = 44), as compared with active smokers (n = 47) and nonexposed nonsmokers (n = 50), to determine whether passive smokers also exhibit altered AA nutriture suggestive of oxidant exposure. Plasma AA concentrations in passive smokers were intermediate between those of active smokers (P = 0.0001) and nonexposed nonsmokers (P = 0.01) despite similar dietary vitamin C intakes. Hypovitaminosis C (< 23 mumol/L) was observed in 24% of active smokers and 12% of passive smokers but not in nonexposed nonsmokers. Reduced plasma AA concentrations were associated with low vitamin C intakes within smoke-exposed populations only. We conclude that chronic smoke exposure, particularly in association with low vitamin C intake, may reduce AA pools in both active and passive smokers.

A study of the potential genotoxicity of cimetidine using human hepatocyte primary cultures: discrepancy from results obtained in rat hepatocytes.

The genotoxicity of cimetidine, a drug widely used in the treatment of peptic ulcer, was examined in human hepatocyte primary cultures. No induction of unscheduled DNA synthesis, as detected by autoradiography, or of DNA fragmentation, as measured by alkaline elution, was seen in metabolically competent human hepatocytes exposed for 20 h to cimetidine concentrations ranging from 0.33 to 9 mM. These findings, which are in contrast with the previously observed capability of cimetidine to induce DNA damage and repair in rat hepatocyte primary cultures, suggest that for some chemicals the rat hepatocyte model might be an inappropriate predictor of potential genotoxic effects in the analogous human cells.

Sentinel node biopsy in the surgical management of breast cancer: experience in a general hospital with a dedicated surgical team.

The aims of this study were to analyse the feasibility and accuracy of the sentinel lymph node biopsy (SLNB) procedure as performed in a general hospital compared with the literature results; to report on the organizational aspects of planning surgical time with higher accuracy of pathological analysis; and to verify that there is a real advantage of SLNB in the surgical management of breast cancer. From October 1999 to September 2000, 371 consecutive patients with T1-2N0 breast lesions underwent SLNB. The immunoscintigraphic method of sentinel node identification was the main one used, the blue dye method being used only when the lymphoscintigraphic method was unsuccessful in identifying sentinel nodes. SLNB was done under either general or local anaesthesia, depending on how the surgical procedure was organized and clinically planned. SLNB was successful in 99% of these T1-2N0 breast cancer cases, and in 71% no metastases were found in the sentinel node. In 47% of cases with axillary metastasis only the sentinel node was involved. Nodal involvement was not present in any case of microinvasive or in situ carcinoma. In T1 cancers nodal involvement was present in 21%; in T2 cases the corresponding rate reached 51%. The results obtained with the SLNB procedure at Bergamo Hospital are similar to the literature data. When a dedicated surgical team, the nuclear medicine department and the pathology department work together, a general hospital can provide breast cancer patients with appropriate surgical treatment.

Population-based sentinel lymph node biopsy in early invasive breast cancer.

INTRODUCTION: Sentinel lymph node biopsy (SLNB) has been proposed as a reliable method for staging of early invasive breast cancer (EIBC). In the present study we analyse the impact of this procedure when systematically applied to all unselected women of a community-based Breast Cancer Unit (BCU). METHODS: All consecutive women with unifocal cT1-2 (