Performance of six diagnostic tests to screen for Chagas disease in blood banks and prevalence of Trypanosoma cruzi infection among donors with inconclusive serology screening based on the analysis of epidemiological variables

OBJECTIVE: The frequent occurrence of inconclusive serology in blood banks and the absence of a gold standard test for Chagas'disease led us to examine the efficacy of the blood culture test and five commercial tests (ELISA, IIF, HAI, c-ELISA, rec-ELISA) used in screening blood donors for Chagas disease, as well as to investigate the prevalence of Trypanosoma cruzi infection among donors with inconclusive serology screening in respect to some epidemiological variables. METHODS: To obtain estimates of interest we considered a Bayesian latent class model with inclusion of covariates from the logit link. RESULTS: A better performance was observed with some categories of epidemiological variables. In addition, all pairs of tests (excluding the blood culture test) presented as good alternatives for both screening (sensitivity > 99.96% in parallel testing) and for confirmation (specificity > 99.93% in serial testing) of Chagas disease. The prevalence of 13.30% observed in the stratum of donors with inconclusive serology, means that probably most of these are non-reactive serology. In addition, depending on the level of specific epidemiological variables, the absence of infection can be predicted with a probability of 100% in this group from the pairs of tests using parallel testing. CONCLUSION: The epidemiological variables can lead to improved test results and thus assist in the clarification of inconclusive serology screening results. Moreover, all combinations of pairs using the five commercial tests are good alternatives to confirm results.

Urediospores of rust fungi are ice nucleation active at > -10°C and harbor ice nucleation active bacteria

Various features of the biology of the rust fungi and of the epidemiology of the plant diseases they cause illustrate the important role of rainfall in their life history. Based on this insight we have characterized the ice nucleation activity (INA) of the aerially disseminated spores (urediospores) of this group of fungi. Urediospores of this obligate plant parasite were collected from natural infections of 7 species of weeds in France, from coffee in Brazil and from field and greenhouse-grown wheat in France, the USA, Turkey and Syria. Immersion freezing was used to determine freezing onset temperatures and the abundance of ice nuclei in suspensions of washed spores. Microbiological analyses of spores from France, the USA and Brazil, and subsequent tests of the ice nucleation activity of the bacteria associated with spores were deployed to quantify the contribution of bacteria to the ice nucleation activity of the spores. All samples of spores were ice nucleation active, having freezing onset temperatures as high as −4 °C. Spores in most of the samples carried cells of ice nucleation-active strains of the bacterium Pseudomonas syringae (at rates of less than 1 bacterial cell per 100 urediospores), but bacterial INA accounted for only a small fraction of the INA observed in spore suspensions. Changes in the INA of spore suspensions after treatment with lysozyme suggest that the INA of urediospores involves a polysaccharide. Based on data from the literature, we have estimated the concentrations of urediospores in air at cloud height and in rainfall. These quantities are very similar to those reported for other biological ice nucleators in these same substrates. However, at cloud level convective activity leads to widely varying concentrations of particles of surface origin, so that mean concentrations can underestimate their possible effects on clouds. We propose that spatial and temporal concentrations of biological ice nucleators active at temperatures > −10 °C and the specific conditions under which they can influence cloud glaciation need to be further evaluated so as to understand how evolutionary processes could have positively selected for INA.

Use of different commercial Artemia sp. Enrichements in Octopus Vulgaris, paralarval rearing at initial planctonic phase: effects on growth, survival and fatty acid composition of the paralarvae

[EN] Octopus "paralarvae", are planktonic, swim actively and have high metabolic rates, requiring large quantities of live prey of adequate motility and nutritional quality ( Iglesias et al., 2000; Navarro and Villanueva, 2000, 2003). During the planktonic phase, they undergo strong morphological changes, after which the octopuses start settling to the bottom. The potential of Octopus vulgaris as candiadate for diversification of marine aquacultures are mainly due to its high food conversion rate and fast growth.( Iglesias et al 2006). Despite the research effort taken until now, paralarval rearing of O. vulgaris still suffers high mortalities which limited the industrial culture of this species. The main problems in the paralarval rearing stages are the high mortality rates and poor growth. These are attributed to the lack of standardized culture techniques and nutritional deficiencies in the diet of paralarvae, especially in n-3 highly unsaturated fatty acids (n-3 HUFA). The objective if this experience was to test different commercial live prey enrichment to improve nutritional quality of the artemia.

Harmful algae and their potential impacts on the coastal ecosystem: growth and toxin production dynamics

The main goal of the present thesis was to study some harmful algal species which cause blooms in Italian coastal waters, leading to consequences for human health, coastal ecosystem, fishery and tourism. In particular, in the first part of this thesis the toxicity of Adriatic strains of the raphidophyte Fibrocapsa japonica was investigated. Despite several hypotheses have been proposed for the toxic mechanism of the raphidophytes, especially for the species Chattonella antiqua and C. marina, which have been studied more extensively, just a few studies on the toxic effects of these species for different organisms were reported. Moreover, a careful reading of the literature evidenced as any ichthyotoxic events reported worldwide can be linked to F. japonica blooms. Although recently several studies were performed on F. japonica strains from the USA, Japan, Australia, New Zealand, the Netherlands, Germany, and France in order to characterize their growth and toxicity features, the work reported in this thesis results one of the first investigation on the toxic effects of F. japonica for different organisms, such as bacteria, crustaceans and fish. Mortality effects, together with haemolysis of fish erythrocytes, probably due to the relatively high amount of PUFAs produced by this species, were observed. Mortality for fish, however, was reported only at a high cell density and after a long exposition period (9-10 days); moreover a significant increase of H2O2 obtained in the tanks where sea basses were exposed to F. japonica was also relevant. This result may justify the absence of ichthyotoxic events in the Italian coasts, despite F. japonica blooms detected in these areas were characterized by high cell densities. This work reports also a first complete characterization of the fatty acids produced and extracellularly released by the Adriatic F. japonica, and results were also compared with the fatty acid profile of other strains. The absence of known brevetoxins in F. japonica algal extracts was also highlighted, leading to the hypothesis that the toxicity of F. japonica may be due to a synergic effect of PUFAs and ROS. Another microalgae that was studied in this thesis is the benthic dinoflagellate Ostreopsis cf. ovata. This species was investigated with the aim to investigate the effect of environmental parameters on its growth and toxicity. O. cf. ovata, in fact, shows different blooming periods along the Italian coasts and even the reported toxic effects are variable. The results of this work confirmed the high variability in the growth dynamic and toxin content of several Italian strains which were isolated in recent years along the Adriatic and Tyrrhenian Seas. Moreover, the effects of temperature and salinity on the behaviour of the different isolates are in good agreement with the results obtained from field surveys, which evidence as the environmental parameters are important factors modulating O. cf. ovata proliferation. Another relevant result that was highlighted is the anomaly in the production of palytoxin-like compounds reported by one of the studied isolate, in particular the one isolated in 2008 in Ancona (Adriatic Sea). Only this strain reported the absence of two (ovatoxin-b and –c) of the five ovatoxins so far known in the toxin profile and a different relative abundance of the other toxins. The last aspect that was studied in this thesis regards the toxin biosythesis. In fact, toxins produced (palytoxin-like compounds) or supposed to be produced (brevetoxin-like compounds) by O. cf. ovata and F. japonica, respectively, are polyketides, which are highly oxygenated compounds synthesized by complex enzymes known as polyketide synthase (PKS) enzymes. These enzymes are multi-domain complexes that structurally and functionally resemble the fatty acid synthases (FASs). This work reports the first study of PKS proteins in the dinoflagellates O. cf. ovata, C. monotis and in the raphidophyte F. japonica. For the first time some PKSs were identified in these species, confirming the presence of PKS proteins predicted by the in silico translation of the transcripts found in K. brevis also in other species. The identification of O. cf. ovata PKSs and the localization of the palytoxin-like compounds produced by this dinoflagellate in a similar location (chloroplast) as that observed for other dinoflagellate and cyanobacterial toxins provides some indication that these proteins may be involved in polyketide biosynthesis. However, their potential function as fatty acid synthases cannot be ruled out, as plant fatty acid synthesis also occurs within chloroplasts. This last hypothesis is also supported by the fact that in all the investigated species, and in particular in F. japonica, PKS proteins were present. Therefore, these results provide an important contribution to the study of the polyketides and of the involvement of PKS proteins in the toxin biosynthesis.

Isolation and characterization of a manganese oxidizing bacterium from the Mediterranean marine sponge Suberites domuncula

In the present study of sponge-bacterial association, the presence of a marine bacterium which has not seen to be associated previously with the Mediterranean sponge Suberites domuncula was investigated. The marine sponge S. domuncula was chosen as the subject of investigation, for the identification of potential symbiotic microorganisms, since it can be kept under controlled laboratory conditions for over five years. By the use of specialized media assisting in the growth of a metal oxidizing bacterium, the manganese oxidizing bacterium was isolated from the surface of the marine sponge. The bacterium so isolated was characterized for its growth characteristics by microbiological and biochemical techniques, a detailed analysis of which showed that the bacterium followed a life cycle where the culture showed the presence of spore forming bacteria. This was correlated to the manganese oxidation activity of the bacteria and it was found that both stages are interdependent.The action of the protein responsible for carrying out the manganese (Mn) oxidation was studied by an in-gel oxidation assay, and the presence of a multi copper oxidase was confirmed by the use of copper chelators in the buffer. In parallel the effect of addition of copper was observed on the manganese oxidation by the bacteria thus supporting the observations. The manganese oxidation reaction by the bacteria was determined in the culture medium and on the surface of the cells, and it could be concluded that the oxidation was facilitated by the presence of the polysaccharides and proteins on the surface of the cells.Thus the presence of a bacterium capable of oxidizing the manganese from the surroundings was confirmed to be symbiotically associated with the marine sponge S. domuncula by monitoring its growth in axenic cultures. The reasons behind this association were studied.This bacterium displays a crucial role in the physiology/metabolism of the sponge by acting as a reversible Mn store in S. domuncula. According to this view, the presence of SubDo-03 bacteria is required as a protection against higher, toxic concentrations of Mn in the environment; manganese (II) after undergoing oxidation to manganese (IV), becomes an insoluble ion. Since only minute levels of manganese exist in the surrounding seawater a substantial accumulation of manganese has to arise, or a release by the bacterial-precipitated manganese (IV) is implicated to maintain the reversible balance. The other possible benefits provided by the bacterial association to the sponge could be in preventing cellular oxygen toxicity, help in nutrient scavenging and detoxification.

Circulating Fibrocytes, their role in renal fibrosis and molecular pathways involved. Possible biomarkers of fibrogenesis in chronic kidney disease

Circulating Fibrocytes (CFs) are bone marrow-derived mesenchymal progenitor cells that express a similar pattern of surface markers related to leukocytes, hematopoietic progenitor cells and fibroblasts. CFs precursor display an ability to differentiate into fibroblasts and Myofibroblasts, as well as adipocytes. Fibrocytes have been shown to contribute to tissue fibrosis in the end-stage renal disease (ESRD), as well as in other fibrotic diseases, leading to fibrogenic process in other organs including lung, cardiac, gut and liver. This evidence has been confirmed by several experimental proofs in mice models of kidney injury. In the present study, we developed a protocol for the study of CFs, by using peripheral blood monocytes cells (PBMCs) samples collected from healthy human volunteers. Thanks to a flow cytometry method, in vitro culture assays and the gene expression assays, we are able to study and characterize this CFs population. Moreover, results confirmed that these approaches are reliable and reproducible for the investigation of the circulating fibrocytes population in whole blood samples. Our final aim is to confirm the presence of a correlation between the renal fibrosis progression, and the different circulating fibrocyte levels in Chronic Kidney Disease (CKD) patients. Thanks to a protocol study presented and accepted by the Ethic Committee we are continuing the study of CFs induction in a cohort of sixty patients affected by CKD, divided in three distinct groups for different glomerular filtration rate (GFR) levels, plus a control group of thirty healthy subjects. Ongoing experiments will determine whether circulating fibrocytes represent novel biomarkers for the study of CKD progression, in the early and late phases of this disease.

Engineering in Transition. Approaches, strategies and technologies for implementing system innovation towards sustainability

With the aim to provide people with sustainable options, engineers are ethically required to hold the safety, health and welfare of the public paramount and to satisfy society's need for sustainable development. The global crisis and related sustainability challenges are calling for a fundamental change in culture, structures and practices. Sustainability Transitions (ST) have been recognized as promising frameworks for radical system innovation towards sustainability. In order to enhance the effectiveness of transformative processes, both the adoption of a transdisciplinary approach and the experimentation of practices are crucial. The evolution of approaches towards ST provides a series of inspiring cases which allow to identify advances in making sustainability transitions happen. In this framework, the thesis has emphasized the role of Transition Engineering (TE). TE adopts a transdisciplinary approach for engineering to face the sustainability challenges and address the risks of un-sustainability. With this purpose, a definition of Transition Technologies is provided as a valid instruments to contribute to ST. In the empirical section, several transition initiatives have been analysed especially at the urban level. As a consequence, the model of living-lab of sustainability has crucially emerged. Living-labs are environments in which innovative technologies and services are co-created with users active participation. In this framework, university can play a key role as learning organization. The core of the thesis has concerned the experimental application of transition approach within the School of Engineering and Architecture of University of Bologna at Terracini Campus. The final vision is to realize a living-lab of sustainability. Particularly, a Transition Team has been established and several transition experiments have been conducted. The final result is not only the improvement of sustainability and resilience of the Terracini Campus, but the demonstration that university can generate solutions and strategies that tackle the complex, dynamic factors fuelling the global crisis.

Escherichia coli variants in periprosthetic joint infection: diagnostic challenges with sessile bacteria and sonication

The diagnostic yield of prosthetic joint-associated infection is hampered by the phenotypic change of bacteria into a sessile and resistant form, also called biofilm. With sonication, adherent bacteria can be dislodged from the prosthesis. Species identification may be difficult because of their variations in phenotypic appearance and biochemical reaction. We have studied the phenotypic, genotypic, and biochemical properties of Escherichia coli variants isolated from a periprosthetic joint infection. The strains were collected from synovial fluid, periprosthetic tissue, and fluid from the explanted and sonicated prosthesis. Isolates from synovial fluid revealed a normal phenotype, whereas a few variants from periprosthetic tissue and all isolates from sonication fluid showed different morphological features (including small-colony variants). All isolates from sonication fluid were beta-galactosidase negative and nonmotile; most were indole negative. Because of further variations in biochemical properties, species identification was false or not possible in 50% of the isolates included in this study. In contrast to normal phenotypes, variants were resistant to aminoglycosides. Typing of the isolates using pulsed-field gel electrophoresis yielded nonidentical banding patterns, but all strains were assigned to the same clonal origin when compared with 207 unrelated E. coli isolates. The bacteria were repeatedly passaged on culture media and reanalyzed. Thereafter, most variants reverted to normal phenotype and regained their motility and certain biochemical properties. In addition, some variants displayed aminoglycoside susceptibility after reversion. Sonication of an explanted prosthesis allows insight into the lifestyle of bacteria in biofilms. Since sonication fluid also reveals dislodged sessile forms, species identification of such variants may be misleading.

Analysis of the erosive effect of different dietary substances and medications

Excessive consumption of acidic drinks and foods contributes to tooth erosion. The aims of the present in vitro study were twofold: (1) to assess the erosive potential of different dietary substances and medications; (2) to determine the chemical properties with an impact on the erosive potential. We selected sixty agents: soft drinks, an energy drink, sports drinks, alcoholic drinks, juice, fruit, mineral water, yogurt, tea, coffee, salad dressing and medications. The erosive potential of the tested agents was quantified as the changes in surface hardness (ΔSH) of enamel specimens within the first 2 min (ΔSH2-0 = SH2 min - SHbaseline) and the second 2 min exposure (ΔSH4-2 = SH4 min - SH2 min). To characterise these agents, various chemical properties, e.g. pH, concentrations of Ca, Pi and F, titratable acidity to pH 7·0 and buffering capacity at the original pH value (β), as well as degree of saturation (pK - pI) with respect to hydroxyapatite (HAP) and fluorapatite (FAP), were determined. Erosive challenge caused a statistically significant reduction in SH for all agents except for coffee, some medications and alcoholic drinks, and non-flavoured mineral waters, teas and yogurts (P < 0·01). By multiple linear regression analysis, 52 % of the variation in ΔSH after 2 min and 61 % after 4 min immersion were explained by pH, β and concentrations of F and Ca (P < 0·05). pH was the variable with the highest impact in multiple regression and bivariate correlation analyses. Furthermore, a high bivariate correlation was also obtained between (pK - pI)HAP, (pK - pI)FAP and ΔSH.

General Protocol for the Culture of Cells on Plasma-Coated Electrospun Scaffolds

As opposed to culture on standard tissue-treated plastic, cell culture on three-dimensional scaffolds impedes additional challenges with respect to substrate preparation, cell seeding, culture maintenance, and analysis. We herewith present a general route for the culture of primary cells, differentiated cells, or stem cells on plasma-coated, electrospun scaffolds. We describe a method to prepare and fix the scaffolds in culture wells and discuss a convenient method for cell seeding and subsequent analysis by scanning electron microscopy or immunohistology.

A Transformation of the Philosophical Thought of West European Liberalism in the Multi-National Central and South-Eastern European Milieu

This work was devoted to individual child development. Psychogenetic research has emphasised the importance of social factors in children's intellectual development and two social factors are looked at here, family size and birth order. The effect of the formal parameters is, however, very unstable and they should therefore be considered together with certain informal factors. Of these, parental educational style, which is an expression of national traditions at the family level, is of particular interest. Educational style is culture-dependent and only a comparative cross-cultural study can reveal the real mechanisms through which educational style influences children's intellectual and personality development. Dumitrascu carried out an experimental cross-cultural study dedicated to the effects of family environment on child intellectual development. This involved three distinct populations, each of which has a distinct status in their geographical area, namely Romanians, Romanies from Romania, and Russians from the Republic of Moldova. It showed a significant difference between child intelligence in those from large families and in only children, with a huge gap in the case of Romany children. This suggests that the simultaneous action of several negative factors (low socio-economic status, large family size, socio-cultural isolation of the population) may delay a child's development. Subjected to such a precarious environment, Romany children do not seek self-realisation but rather struggle to overcome hardship and the majority remain outside civilisation. Unfortunately, adult Romanies rarely show concern about their children's successful social integration, placing no value on the school as a major socialising tool. This leads to the conclusion that a major effort is needed to help Romanies' social integration.

The Social Desintegration of an Ethnical German Group from Romanian Village Community Comparative with a Romanian Group and Hungarian Group from some Village Their Integration into the German Society

Zarna is witness to the disappearance of the Swabian community in Santana, a process which seems to date back to before the major changes in Central and Eastern Europe. His project showed how a strong German ethnic community, formed more than 250 years ago, has virtually disappeared from the village of Santana (Romania). Zarna presents the causes leading to the loss of their ethnic identity, of their culture, traditions and of the collective reproduction of their ethnicity, although the last Swabians remaining in the village have preserved their individual identity and not let themselves be assimilated. The policy of the former communist regime is not sufficient to explain the decline of the German ethnic group, nor is the present international context with its varying effects on the form and reproduction of their ethnic identity. Zarna has analysed the origins of the Swabian community, its development, historical changes (both desired and imposed) and the disappearance of elements that determined their German culture and their pride in being German. The Germans have demobilised more rapidly than other ethnic groups in Romania, partly because of Germany's pro-emigration policy over the last two decades. Many of the emigrants were however, poorly prepared for emigration and have not been able to recreate the prosperous financial situation which they left. The prevalent feeling among those interviewed was disappointment and this increases with age and education.

Assessing the expression of enterotoxigenic Escherichia coli-specific surface antigens in recombinant strains by transmission electron microscopy and immunolabeling

Infections with enterotoxigenic Escherichia coli (ETEC) are a major cause of travelers' diarrhea worldwide. Colonization of the small intestine mucosa is dependent on specific colonization factor antigens (CFA) and coli surface (CS) antigens. CFA/1, CS3, and CS6 are the most prevalent fimbrial antigens found in clinical isolates. The goal of our study was to visualize the morphology of CS3 and CS6 fimbriae in wild-type and recombinant E. coli strains by means of transmission electron microscopy in conjunction with negative staining and immunolabeling. Corresponding ETEC genes were cloned into E. coli K12 strain DH10B. Expression of fimbriae was dependent on culture conditions and sample handling. Specific immunolabeling of fimbriae unequivocally demonstrated the presence of all types of surface antigens investigated. Negative staining was effective in revealing CS3 but not CS6. In addition, this technique clearly demonstrated differences in the morphology of genetically and immunologically identical CS3 surface antigens in wild-type and recombinant strains. This paper provides a basis for the assessment of recombinant vaccines.

In vitro effects of thiazolides on Giardia lamblia WB clone C6 cultured axenically and in coculture with Caco2 cells

The thiazolides represent a novel class of anti-infective drugs, with the nitrothiazole nitazoxanide [2-acetolyloxy-N-(5-nitro 2-thiazolyl) benzamide] (NTZ) as the parent compound. NTZ exhibits a broad spectrum of activities against a wide variety of helminths, protozoa, and enteric bacteria infecting animals and humans. In vivo, NTZ is rapidly deacetylated to tizoxanide (TIZ), which exhibits similar activities. We have here comparatively investigated the in vitro effects of NTZ, TIZ, a number of other modified thiazolides, and metronidazole (MTZ) on Giardia lamblia trophozoites grown under axenic culture conditions and in coculture with the human cancer colon cell line Caco2. The modifications of the thiazolides included, on one hand, the replacement of the nitro group on the thiazole ring with a bromide, and, on the other hand, the differential positioning of methyl groups on the benzene ring. Of seven compounds with a bromo instead of a nitro group, only one, RM4820, showed moderate inhibition of Giardia proliferation in axenic culture, but not in coculture with Caco2 cells, with a 50% inhibitory concentration (IC50) of 18.8 microM; in comparison, NTZ and tizoxanide had IC50s of 2.4 microM, and MTZ had an IC50 of 7.8 microM. Moreover, the methylation or carboxylation of the benzene ring at position 3 resulted in a significant decrease of activity, and methylation at position 5 completely abrogated the antiparasitic effect of the nitrothiazole compound. Trophozoites treated with NTZ showed distinct lesions on the ventral disk as soon as 2 to 3 h after treatment, whereas treatment with metronidazole resulted in severe damage to the dorsal surface membrane at later time points.