Diagnostic value of cone beam computed tomography for root canal morphology assessment - a micro-CT based comparison.

OBJECTIVES: The aim of this study was to assess cone beam computed tomography (CBCT) as a root canal anatomy diagnostic tool by comparison with micro-CT gold-standard. MATERIALS AND METHODS: 216 two-rooted mandibular molars were first scanned in a CBCT device (200 µm voxel size) and posteriorly in a micro-CT scanner (19.61 µm). The volumes were sequentially screened to classify main root canal anatomy according to Vertucci classification, and for the presence of lateral canals and apical deltas, in both mesial and distal roots. RESULTS: Both methods revealed a higher prevalence of Vertucci Type II and IV in the mesial root, and Vertucci Type I in the distal root. The percentage of agreement for main root canal anatomy classification between CBCT and micro-CT scores was high (85.2%). CONCLUSION: Sensibility to detect both lateral canals and apical deltas with CBCT was low. These results attest to the fact that minor anatomical changes might be difficult to identify with CBCT imaging, hampering its diagnostic value.

Insights into the transcriptional regulation of poorly characterized alcohol acetyltransferase-encoding genes (HgAATs) shed light into the production of acetate esters in the wine yeast Hanseniaspora guilliermondii.

Hanseniaspora guilliermondii is a well-recognized producer of acetate esters associated with fruity and floral aromas. The molecular mechanisms underneath this production or the environmental factors modulating it remain unknown. Herein, we found that, unlike Saccharomyces cerevisiae, H. guilliermondii over-produces acetate esters and higher alcohols at low carbon-to-assimilable nitrogen (C:N) ratios, with the highest titers being obtained in the amino acid-enriched medium YPD. The evidences gathered support a model in which the strict preference of H. guilliermondii for amino acids as nitrogen sources results in a channeling of keto-acids obtained after transamination to higher alcohols and acetate esters. This higher production was accompanied by higher expression of the four HgAATs, genes, recently proposed to encode alcohol acetyl transferases. In silico analyses of these HgAat's reveal that they harbor conserved AATs motifs, albeit radical substitutions were identified that might result in different kinetic properties. Close homologues of HgAat2, HgAat3, and HgAat4 were only found in members of Hanseniaspora genus and phylogenetic reconstruction shows that these constitute a distinct family of Aat's. These results advance the exploration of H. guilliermondii as a bio-flavoring agent providing important insights to guide future strategies for strain engineering and media manipulation that can enhance production of aromatic volatiles.

Multimethod Assessment of the Cyclic Fatigue Strength of ProGlider, Edge Glide Path and R-Pilot Endodontic Instruments.

BACKGROUND: The aim of this study was to comprehensively evaluate the cyclic fatigue strength of ProGlider, Edge Glide Path, and R-Pilot instruments. METHODS: Sixty-three instruments were submitted to a multimethod evaluation. Their design was analyzed by stereomicroscopy and scanning electron microscopy, including the number of blades, helical angle means, cross-sectional design, surface finishing, and symmetry. Energy-dispersive X-ray spectroscopy was used determine the nickel/titanium elements ratio, and differential scanning calorimetry determined the instruments' phase transformation temperatures. The cyclic fatigue tests were conducted in an artificial canal with a 6 mm radius and 86 degrees of curvature. The Mood's median test and one-way ANOVA were used to determine differences, with the significance level set at 0.05. RESULTS: The ProGlider presented the highest number of blades (n = 21), while R-Pilot had the highest helical angles (26.4°). Differences were noted in the instruments' cross-sections and surface finishing. The ProGlider and R-Pilot showed some similarities regarding the phase transformation temperatures but differed from the Edge Glide Path. All alloys showed an almost equiatomic nickel/titanium ratio. The R-Pilot instruments showed a significantly higher (p < 0.05) time to fracture than both the other files. CONCLUSION: Reciprocating R-Pilot instruments showed a higher cyclic fatigue time to fracture than the ProGlider and Edge Glide Path rotary files.

Machine Learning Techniques Disclose the Combined Effect of Fermentation Conditions on Yeast Mixed-Culture Dynamics and Wine Quality.

The use of yeast starter cultures consisting of a blend of Saccharomyces cerevisiae and non-Saccharomyces yeasts has increased in recent years as a mean to address consumers' demands for diversified wines. However, this strategy is currently limited by the lack of a comprehensive knowledge regarding the factors that determine the balance between the yeast-yeast interactions and their responses triggered in complex environments. Our previous studies demonstrated that the strain Hanseniaspora guilliermondii UTAD222 has potential to be used as an adjunct of S. cerevisiae in the wine industry due to its positive impact on the fruity and floral character of wines. To rationalize the use of this yeast consortium, this study aims to understand the influence of production factors such as sugar and nitrogen levels, fermentation temperature, and the level of co-inoculation of H. guilliermondii UTAD222 in shaping fermentation and wine composition. For that purpose, a Central Composite experimental Design was applied to investigate the combined effects of the four factors on fermentation parameters and metabolites produced. The patterns of variation of the response variables were analyzed using machine learning methods, to describe their clustered behavior and model the evolution of each cluster depending on the experimental conditions. The innovative data analysis methodology adopted goes beyond the traditional univariate approach, being able to incorporate the modularity, heterogeneity, and hierarchy inherent to metabolic systems. In this line, this study provides preliminary data and insights, enabling the development of innovative strategies to increase the aromatic and fermentative potential of H. guilliermondii UTAD222 by modulating temperature and the availability of nitrogen and/or sugars in the medium. Furthermore, the strategy followed gathered knowledge to guide the rational development of mixed blends that can be used to obtain a particular wine style, as a function of fermentation conditions.

3D-printed multisampling holder for microcomputed tomography applied to life and materials science research.

The aim of this work was to design, fabricate, test and validate a 3D-printed multisampling holder for multi-analysis by microcomputed tomography. Different raw materials were scanned by microcomputed tomography. The raw material chosen was used to fabricate the holder by 3D printing. To validate the multisampling holder, five teeth were filled with a high density-material and scanned in two ways: a single and a multisampling scan mode. For each tooth, the root canal filling volume, porosity volume, closed pore volume, and open pore volume were calculated and compared when the same tooth was scanned in the two sampling scan mode. ABSplus P430™ allowed a high transmission value (84.3 %), and then it was the polymeric material selected to fabricate the holder. In a single sampling scan mode, the scan duration for scanning five teeth was 87.42 min, contrasting with 21.51 min for a multisampling scan mode, which scanned five teeth at the same time. The scan duration time and the cost using a multisampling holder represented a reduction of 75 % and the data volume generated represented a reduction of 60 %. Comparing the two scan modes, the results also showed that the difference of root canal filling volume, porosity volume, closed pore volume, and open pore volume was not statistically significant (p > .05). The multisampling holder was validated to do multi-analysis by microcomputed tomography without significant loss of quantitative accuracy data, allowing a reduction in scan duration time, imaging cost, and data storage.

Effect of different manipulations on the physical, chemical and microstructural characteristics of Biodentine.

OBJECTIVE: The water to powder ratio and method of mixing is important for the properties of hydraulic cements. For this purpose a number of clinicians prefer premixed materials. Dental manufacturing companies provide predosed materials, however the manufacturer instructions are not always adhered to. The aim of this research is to investigate physical and chemical alterations of the tricalcium silicate-based cement Biodentine when manipulated according to the manufacturer's instructions (control) or changing the doses and mixing of the material components. METHODS: 6 groups were constituted according to different mixing and dosing of powder and liquid. The hydrated cements were characterized using scanning electron microscopy (SEM), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR). Calcium ion concentration of the leachate was also investigated. Assessment of the physical characteristics included setting time and microhardness. RESULTS: Microstructural differences were visible only in the Biodentine mixed manually with water, in which early hydration rate was also affected, with lower calcium ion release. Increase of Biodentine liquid increased the calcium ion release, but also increased the setting time. Manual manipulation required more liquid (both water and Biodentine liquid) added to the mixture to guarantee a similar consistency to the control. A decrease in setting time was also noted. All groups showed higher values of microhardness at 24 h compared to the freshly set materials. In the freshly set materials, there was an overall decrease in microhardness in all groups when compared to group control, particularly significant when increasing the dosage of Biodentine liquid. SIGNIFICANCE: When mixing Biodentine, altering the mixing procedure in terms of type and amount of liquid added to the powder and mixing device chosen has an effect on the physical, chemical and mechanical characteristics and surface topography of the material, when compared to Biodentine mixed according to the manufacturer's recommendations. Hence, the manufacturer's instructions should be strictly followed.

Characterizing the Potential of the Non-Conventional Yeast Saccharomycodes ludwigii UTAD17 in Winemaking.

Non-Saccharomyces yeasts have received increased attention by researchers and winemakers, due to their particular contributions to the characteristics of wine. In this group, Saccharomycodes ludwigii is one of the less studied species. In the present study, a native S. ludwigii strain, UTAD17 isolated from the Douro wine region was characterized for relevant oenological traits. The genome of UTAD17 was recently sequenced. Its potential use in winemaking was further evaluated by conducting grape-juice fermentations, either in single or in mixed-cultures, with Saccharomyces cerevisiae, following two inoculation strategies (simultaneous and sequential). In a pure culture, S. ludwigii UTAD17 was able to ferment all sugars in a reasonable time without impairing the wine quality, producing low levels of acetic acid and ethyl acetate. The overall effects of S. ludwigii UTAD17 in a mixed-culture fermentation were highly dependent on the inoculation strategy which dictated the dominance of each yeast strain. Wines whose fermentation was governed by S. ludwigii UTAD17 presented low levels of secondary aroma compounds and were chemically distinct from those fermented by S. cerevisiae. Based on these results, a future use of this non-Saccharomyces yeast either in monoculture fermentations or as a co-starter culture with S. cerevisiae for the production of wines with greater expression of the grape varietal character and with flavor diversity could be foreseen.

Development and optimization of a HS-SPME-GC-MS methodology to quantify volatile carbonyl compounds in Port wines.

A method based on headspace solid-phase microextraction (HS-SPME) coupled to gas chromatography-triple quadrupole/mass spectrometry detection (GC-TQ/MS) with a prior derivatization step with O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride (PFBHA) was developed to quantify carbonyl compounds in different categories of Port wines. Optimal extraction conditions were obtained incubating 2 ml of wine with 2.3 g/l of PFBHA for 10 min and extracted during 20 min at 32 °C. The method was validated for 38 carbonyl compounds (alkanals, alkenals, Strecker aldehydes, dialdehydes, ketones and furan aldehydes) with regard to linearity, repeatability, inter and intra-day precision and accuracy, showing that the method is suitable for the determination of carbonyl compounds in wines. Tawny wines with 'indication of age' (10-40 years old) presented the highest levels of some carbonyl compounds, such as propanal, pentanal, hexanal, Strecker aldehydes, diacetyl, methyl glyoxal, 3-pentanone and 2-furfural, whereas Ruby wines were characterized by the highest amounts of some unidentified compounds.

H. guilliermondii impacts growth kinetics and metabolic activity of S. cerevisiae: the role of initial nitrogen concentration.

Non-Saccharomyces yeasts include different species which comprise an ecologically and biochemically diverse group capable of altering fermentation dynamics and wine composition and flavour. In this study, single- and mixed-culture of Hanseniaspora guilliermondii and Saccharomyces cerevisiae were used to ferment natural grape-juice, under two nitrogen regimes. In single-culture the strain H. guilliermondii failed to complete total sugar breakdown even though the nitrogen available has not been a limiting factor of its growth or fermentative activity. In mixed-culture, that strain negatively interfered with the growth and fermentative performance of S. cerevisiae, resulting in lower fermentation rate and longer fermentation length, irrespective of the initial nitrogen concentration. The impact of co-inoculation on the volatile compounds profile was more evident in the wines obtained from DAP-supplemented musts, characterised by increased levels of ethyl and acetate esters, associated with fruity and floral character of wines. Moreover, the levels of fatty acids and sulphur compounds which are responsible for unpleasant odours that depreciate wine sensory quality were significantly lower. Accordingly, data obtained suggests that the strain H. guilliermondii has potential to be used as adjunct of S. cerevisiae in wine industry, although possible interactions with S. cerevisiae still need to be elucidated.

1,3-Propanediol production in a two-step process fermentation from renewable feedstock.

In this work, the production of 1,3-propanediol from glucose and molasses was studied in a two-step process using two recombinant microorganisms. The first step of the process is the conversion of glucose or other sugar into glycerol by the metabolic engineered Saccharomyces cerevisiae strain HC42 adapted to high (>200 g l(-1)) glucose concentrations. The second step, carried out in the same bioreactor, was performed by the engineered strain Clostridium acetobutylicum DG1 (pSPD5) that converts glycerol to 1,3-propanediol. This two-step strategy led to a flexible process, resulting in a 1,3-propanediol production and yield that depended on the initial sugar concentration. Below 56.2 g l(-1) of sugar concentration, cultivation on molasses or glucose showed no significant differences. However, at higher molasses concentrations, glycerol initially produced by yeast could not be totally converted into 1,3-propanediol by C. acetobutylicum and a lower 1,3-propanediol overall yield was observed. In our hand, the best results were obtained with an initial glucose concentration of 103 g l(-1), leading to a final 1,3-propanediol concentration of 25.5 g l(-1), a productivity of 0.16 g l(-1) h(-1) and 1,3-propanediol yields of 0.56 g g(-1) glycerol and 0.24 g g(-1) sugar, which is the highest value reported for a two-step process. For an initial sugar concentration (from molasses) of 56.2 g l(-1), 27.4 g l(-1) of glycerol were produced, leading to 14.6 g l(-1) of 1.3-propanediol and similar values of productivity, 0.15 g l(-1) h(-1), and overall yield, 0.26 g g(-1) sugar.

Relationship between nitrogen content in grapes and volatiles, namely heavy sulphur compounds, in wines.

Ammonium salts were added to white grape musts, before alcoholic fermentation, in order to evaluate their influence on the heavy sulphur compound and aliphatic higher alcohol composition of resulting wines. Six grape musts were used (Trajadura, Pedernã, Loureiro, Azal Branco, Avesso and Alvarinho). Ammonium supplementation of Trajadura and Pedernã grape musts, with the highest nitrogen level, did not influence the content of heavy sulphur compounds and aliphatic higher alcohols in wines; however, the addition of ammonium salts to grape musts with low initial nitrogen content, such as Loureiro, Azal Branco and Avesso, led to a higher production of 1-propanol and a lower production of isoamyl alcohols and sulphur compounds, e.g. S-methyl thioacetate, 2-mercaptoethanol, acetic acid-3-(methylthio)propyl ester, 3-mercapto-1-propanol, 4-(methylthio)-1-butanol, 3-(ethylthio)-1-propanol, 3-methylthiopropionic acid and N-3-(methylthiopropyl)acetamide. For Alvarinho grape must, a decrease in sulphur compound concentrations in wines was only observed for 3-methylthiopropionic acid, acetic acid-3-(methylthio)propyl ester and 2-mercaptoethanol.

Sequential injection kinetic flow assay for monitoring glycerol in a sugar fermentation process by Saccharomyces cerevisiae.

A sequential injection system to monitor glycerol in a Saccharomyces cerevisiae fermentation process was developed. The method relies on the rate of formation of nicotinamide adenine dinucleotide in its reduced form (NADH, measured spectrophotometrically at 340 nm) from the reaction of glycerol with NAD(+) cofactor, catalysed by the enzyme glycerol dehydrogenase present in solution. This procedure enables the determination of glycerol between 0.046 and 0.46 g/l, (corresponding to yeast fermentation samples with concentrations up to 50 g/l) with good repeatability (relative standard deviation for n = 10 lower than 2.2% for three different samples) at a sampling frequency of 25/h. The detection and quantification limits using a miniaturised spectrophotometer were 0.13 and 0.44 mM, respectively. Reagent consumption was of 0.45 mumol NAD(+) and 1.8 microg enzyme per assay, and the waste production was 2.8 ml per determination. Results obtained for samples were in agreement with those obtained with a high-performance liquid chromatography method.

Concepções de loucura em um traçado histórico-cultural: uma articulação com o Construcionismo

O presente artigo, expectador das diferentes concepções de loucura em função de fatores históricos e culturais, visa discorrer sobre estas temáticas, articulando-as com o posicionamento teórico do Construcionismo Social. Esta perspectiva culturalmente orientada preconiza a forte influência dos fatores contextuais, sociais, econômicos e geográficos, destacando, sobre¬tudo, seu caráter pragmático, parcial e relativista. Desta forma, temos o escopo de apresentar algumas considerações sobre o Construcionismo Social e discorrer sobre algumas concepções de loucura. Como objetivo secundário, visamos fragilizar a postura universal na leitura desses fenômenos, ressaltando como ela pode impor limitações aos profissionais de saúde mental e, conse¬quentemente, aos seus pacientes.(AU)

This article, aware of different conceptions of madness by historical and cultural factors, aims to discuss certain issues, articulating them with theoretical position of Social Constructionism. That prospect culturally oriented suggests the strong influence of contextual, social, economic and geographical factors, putting in focus its pragmatic, partial and relativist feature. Thus, we will present some considerations about Social Constructionism, and discuss some conceptions of madness. As a secondary goal, we aim to weaken the universal posture about these phenomena, emphasizing how it can impose limitations on mental health professionals, and thus to patients.(AU)

En este artículo, las expectativas de los diferentes conceptos de la locura a la luz de factores históricos y culturales, tiene por objeto examinar estas cuestiones, la articulación con la posición teórica del Construccionismo Social. El enfoque orientado hacia los defensores de la cultura de la fuerte influencia de factores contextuales, sociales, económicos y geográficos, entre ellos, sobre todo, su carácter pragmático, relativista y parcial. Por lo tanto, nuestro objetivo es presentar algunas consideraciones sobre el construccionismo social y discutir algunas ideas de la locura. Como objetivo secundario, con el objetivo de debilitar la actitud universal en la lectura de estos fenómenos, haciendo hincapié en cómo esto puede imponer limitaciones a los profesionales de salud mental y, en consecuencia, sus pacientes.(AU)

Cet article, les attentes des différents concepts de l’aliénation mentale, à la lumière de facteurs historiques et culturels, a pour but de discuter de ces questions, de les articuler avec la position théorique de la social Construc¬tionism. Les défenseurs de l’approche axée sur la culture de la forte influence de facteurs contextuels, sociaux, économiques et géographiques, y compris par-dessus tout, son caractère pragmatique, partielle et relativiste. Ainsi, nous avons pour objectif de présenter quelques considérations sur le plan social Constructionism et de discuter des idées de folie. Comme objectif secondaire, qui vise à affaiblir l’attitude universelle dans la lecture de ces phénomènes, en mettant l’accent sur la façon d’imposer des restrictions sur les professionnels de la santé mentale et, par conséquent, de leurs patients.(AU)

Concepções de loucura em um traçado histórico-cultural: uma articulação com o Construcionismo / Conceptions of madness in a historical and cultural route: an association with Social Constructionism

O presente artigo, expectador das diferentes concepções de loucura em função de fatores históricos e culturais, visa discorrer sobre estas temáticas, articulando-as com o posicionamento teórico do Construcionismo Social. Esta perspectiva culturalmente orientada preconiza a forte influência dos fatores contextuais, sociais, econômicos e geográficos, destacando, sobre¬tudo, seu caráter pragmático, parcial e relativista. Desta forma, temos o escopo de apresentar algumas considerações sobre o Construcionismo Social e discorrer sobre algumas concepções de loucura. Como objetivo secundário, visamos fragilizar a postura universal na leitura desses fenômenos, ressaltando como ela pode impor limitações aos profissionais de saúde mental e, conse¬quentemente, aos seus pacientes.

This article, aware of different conceptions of madness by historical and cultural factors, aims to discuss certain issues, articulating them with theoretical position of Social Constructionism. That prospect culturally oriented suggests the strong influence of contextual, social, economic and geographical factors, putting in focus its pragmatic, partial and relativist feature. Thus, we will present some considerations about Social Constructionism, and discuss some conceptions of madness. As a secondary goal, we aim to weaken the universal posture about these phenomena, emphasizing how it can impose limitations on mental health professionals, and thus to patients.

En este artículo, las expectativas de los diferentes conceptos de la locura a la luz de factores históricos y culturales, tiene por objeto examinar estas cuestiones, la articulación con la posición teórica del Construccionismo Social. El enfoque orientado hacia los defensores de la cultura de la fuerte influencia de factores contextuales, sociales, económicos y geográficos, entre ellos, sobre todo, su carácter pragmático, relativista y parcial. Por lo tanto, nuestro objetivo es presentar algunas consideraciones sobre el construccionismo social y discutir algunas ideas de la locura. Como objetivo secundario, con el objetivo de debilitar la actitud universal en la lectura de estos fenómenos, haciendo hincapié en cómo esto puede imponer limitaciones a los profesionales de salud mental y, en consecuencia, sus pacientes.

Cet article, les attentes des différents concepts de l’aliénation mentale, à la lumière de facteurs historiques et culturels, a pour but de discuter de ces questions, de les articuler avec la position théorique de la social Construc¬tionism. Les défenseurs de l’approche axée sur la culture de la forte influence de facteurs contextuels, sociaux, économiques et géographiques, y compris par-dessus tout, son caractère pragmatique, partielle et relativiste. Ainsi, nous avons pour objectif de présenter quelques considérations sur le plan social Constructionism et de discuter des idées de folie. Comme objectif secondaire, qui vise à affaiblir l’attitude universelle dans la lecture de ces phénomènes, en mettant l’accent sur la façon d’imposer des restrictions sur les professionnels de la santé mentale et, par conséquent, de leurs patients.

Considerações sobre as múltiplas formas de violência doméstica / Considerations on the many forms of domestic violence

As estatísticas referentes à violência intrafamiliar/doméstica têm estado em constante progressão, constituindo um problema social de relevantes consequências. Com o escopo maior de pôr em destaque um tema tão recorrente em debates multidisciplinares, discute-se os três principais tipos de violência doméstica (a física, a psicológica e a sexual), ressaltando as conseqüências específicas ao agredido e aspectos peculiares em casa forma de agressão. Foram tecidas, também, algumas considerações sobre questões pertinentes e algumas possibilidades para os profissionais que com ela se defrontarem. Com a discussão, conclui-se que a violência doméstica apresenta-se como um fenômeno complexo, cuja ajuda e cujo reconhecimento de todos aqueles que puderem identificar tais abusos são imperiosos no combate a ele. Assim, reitera-se o caráter obscuro decorrente da sua prática, que deve ser combatido não apenas pela parte vitimada, mas por toda a sociedade que, naturalmente, busca ser sadia(AU)

The statistics relating to domestic violence have been steadily increasing, constituting a social problem of relevant consequences. In order to emphasize such a recurrent theme in multidisciplinary discussions, this article discusses the three main tYpes if domestic violence (physical, psychological and sexual), highlighting the specific consequences and peculiar aspects of each form of aggression. Comments were also made about relevant issues and some opportunities for prrfessionals that have to deal with them. With this discussion, the study concludes that domestic violence presents itself as a complex phenomenon, and that the help and recognition of all those who can identify such abuses are essential infighting them. Thus, it refocuses on the hidden nature if domestic violence, which must be fought not only by the victim, but by society as a whole, which naturally seeks to be healthy(AU)

A metabolic and genomic study of engineered Saccharomyces cerevisiae strains for high glycerol production.

Towards a global objective to produce chemical derivatives by microbial processes, this work dealt with a metabolic engineering of the yeast Saccharomyces cerevisiae for glycerol production. To accomplish this goal, overexpression of GPD1 was introduced in a tpi1delta mutant defective in triose phosphate isomerase. This strategy alleviated the inositol-less phenotype of this mutant, by reducing the levels of dihydroxyacetone phosphate and glycerol-3-P, two potent inhibitors of myo-inositol synthase that catalyzes the formation of inositol-6-phosphate from glucose-6-phosphate. Further deletion of ADH1 and overexpression of ALD3, encoding, respectively, the major NAD+-dependent alcohol dehydrogenase and a cytosolic NAD+-dependent aldehyde dehydrogenase yielded a yeast strain able to produce 0.46 g glycerol (g glucose)(-1) at a maximal rate of 3.1 mmol (g dry mass)(-1) h(-1) in aerated batch cultures. At the metabolic level, this genetic strategy shifted the flux control coefficient of the pathway to the level of the glycerol efflux, with a consequent intracellular accumulation of glycerol that could be partially reduced by the overproduction of glycerol exporter encoded by FPS1. At the transcriptomic level, this metabolic reprogramming brought about the upregulation of genes encoding NAD+/NADP+ binding proteins, a partial derepression of genes coding for TCA cycle and respiratory enzymes, and a downregulation of genes implicated in protein biosynthesis and ribosome biogenesis. Altogether, these metabolic and molecular alterations stand for major hurdles that may represent potential targets for further optimizing glycerol production in yeast.

Microbial conversion of glycerol to 1,3-propanediol: physiological comparison of a natural producer, Clostridium butyricum VPI 3266, and an engineered strain, Clostridium acetobutylicum DG1(pSPD5).

Clostridium acetobutylicum is not able to grow on glycerol as the sole carbon source since it cannot reoxidize the excess of NADH generated by glycerol catabolism. Nevertheless, when the pSPD5 plasmid, carrying the NADH-consuming 1,3-propanediol pathway from C. butyricum VPI 3266, was introduced into C. acetobutylicum DG1, growth on glycerol was achieved, and 1,3-propanediol was produced. In order to compare the physiological behavior of the recombinant C. acetobutylicum DG1(pSPD5) strain with that of the natural 1,3-propanediol producer C. butyricum VPI 3266, both strains were grown in chemostat cultures with glycerol as the sole carbon source. The same "global behavior" was observed for both strains: 1,3-propanediol was the main fermentation product, and the qH2 flux was very low. However, when looking at key intracellular enzyme levels, significant differences were observed. Firstly, the pathway for glycerol oxidation was different: C. butyricum uses a glycerol dehydrogenase and a dihydroxyacetone kinase, while C. acetobutylicum uses a glycerol kinase and a glycerol-3-phosphate dehydrogenase. Secondly, the electron flow is differentially regulated: (i) in C. butyricum VPI 3266, the in vitro hydrogenase activity is 10-fold lower than that in C. acetobutylicum DG1(pSPD5), and (ii) while the ferredoxin-NAD+ reductase activity is high and the NADH-ferredoxin reductase activity is low in C. acetobutylicum DG1(pSPD5), the reverse is observed for C. butyricum VPI 3266. Thirdly, lactate dehydrogenase activity is only detected in the C. acetobutylicum DG1(pSPD5) culture, explaining why this microorganism produces lactate.

Alcohols, esters and heavy sulphur compounds production by pure and mixed cultures of apiculate wine yeasts.

Strains of Hanseniaspora uvarum, Hanseniaspora guilliermondii and Saccharomyces cerevisiae were used as pure or mixed starter cultures in commercial medium, in order to compare their kinetic parameters and fermentation patterns. In pure and mixed cultures, yeasts presented similar ethanol yield and productivity. Pure cultures of H. uvarum and S. cerevisiae showed a specific growth rate of 0.38 h(-1); however, this value decreased when these yeasts were grown in mixed cultures with H. guilliermondii. The specific growth rate of pure cultures of H. guilliermondii was 0.41 h(-1) and was not affected by growth of other yeasts. H. guilliermondii was found to be the best producer of 2-phenylethyl acetate and 2-phenylethanol in both pure and mixed cultures. In pure cultures, H. uvarum led to the highest contents of heavy sulphur compounds, but H. guilliermondii and S. cerevisiae produced similar levels of methionol and 2-methyltetrahydrothiophen-3-one. Growth of apiculate yeasts in mixed cultures with S. cerevisiae led to amounts of 3-methylthiopropionic acid, acetic acid-3-(methylthio)propyl ester and 2-methyltetrahydrothiophen-3-one similar to those obtained in a pure culture of S. cerevisiae; however, growth of apiculate yeasts increased methionol contents of fermented media.

Metabolic engineering of Clostridium acetobutylicum for the industrial production of 1,3-propanediol from glycerol.

Clostridium butyricum is to our knowledge the best natural 1,3-propanediol producer from glycerol and the only microorganism identified so far to use a coenzyme B12-independent glycerol dehydratase. However, to develop an economical process of 1,3-propanediol production, it would be necessary to improve the strain by a metabolic engineering approach. Unfortunately, no genetic tools are currently available for C. butyricum and all our efforts to develop them have been so far unsuccessful. To obtain a better "vitamin B12-free" biological process, we developed a metabolic engineering strategy with Clostridium acetobutylicum. The 1,3-propanediol pathway from C. butyricum was introduced on a plasmid in several mutants of C. acetobutylicum altered in product formation. The DG1(pSPD5) recombinant strain was the most efficient strain and was further characterized from a physiological and biotechnological point of view. Chemostat cultures of this strain grown on glucose alone produced only acids (acetate, butyrate and lactate) and a high level of hydrogen. In contrast, when glycerol was metabolized in chemostat culture, 1,3-propanediol became the major product, the specific rate of acid formation decreased and a very low level of hydrogen was observed. In a fed-batch culture, the DG1(pSPD5) strain was able to produce 1,3-propanediol at a higher concentration (1104 mM) and productivity than the natural producer C. butyricum VPI 3266. Furthermore, this strain was also successfully used for very long term continuous production of 1,3-propanediol at high volumetric productivity (3 g L-1 h-1) and titer (788 mM).

Avaliação funcional do paciente respirador bucal / Functional evaluation of mouth breathing patient

O objetivo deste artigo foi de salientar a necessidade de diagnóstico e tratamento precoce do paciente com obstrução respiratória, visto que esta repercute no crescimento e desenvolvimento da criança de forma muito negativa. Foi realizada uma pesquisa através do exame clínico de 60 pacientes de uma clínica particular, utilizando-se uma ficha de avaliação funcional