Single nucleotide polymorphisms associated with wine fermentation and adaptation to nitrogen limitation in wild and domesticated yeast strains.

For more than 20 years, Saccharomyces cerevisiae has served as a model organism for genetic studies and molecular biology, as well as a platform for biotechnology (e.g., wine production). One of the important ecological niches of this yeast that has been extensively studied is wine fermentation, a complex microbiological process in which S. cerevisiae faces various stresses such as limited availability of nitrogen. Nitrogen deficiencies in grape juice impair fermentation rate and yeast biomass production, leading to sluggish or stuck fermentations, resulting in considerable economic losses for the wine industry. In the present work, we took advantage of the "1002 Yeast Genomes Project" population, the most complete catalogue of the genetic variation in the species and a powerful resource for genotype-phenotype correlations, to study the adaptation to nitrogen limitation in wild and domesticated yeast strains in the context of wine fermentation. We found that wild and domesticated yeast strains have different adaptations to nitrogen limitation, corroborating their different evolutionary trajectories. Using a combination of state-of-the-art bioinformatic (GWAS) and molecular biology (CRISPR-Cas9) methodologies, we validated that PNP1, RRT5 and PDR12 are implicated in wine fermentation, where RRT5 and PDR12 are also involved in yeast adaptation to nitrogen limitation. In addition, we validated SNPs in these genes leading to differences in fermentative capacities and adaptation to nitrogen limitation. Altogether, the mapped genetic variants have potential applications for the genetic improvement of industrial yeast strains.

Yeast as a biological platform for vitamin D production: A promising alternative to help reduce vitamin D deficiency in humans.

Vitamin D is an important human hormone, known primarily to be involved in the intestinal absorption of calcium and phosphate, but it is also involved in various nonskeletal processes (molecular, cellular, immune, and neuronal). One of the main health problems nowadays is the vitamin D deficiency of the human population due to lack of sun exposure, with estimates of one billion people worldwide with vitamin D deficiency, and the consequent need for clinical intervention (i.e., prescription of pharmacological vitamin D supplements). An alternative to reduce vitamin D deficiency is to produce good dietary sources of it, a scenario in which the yeast Saccharomyces cerevisiae seems to be a promising alternative. This review focuses on the potential use of yeast as a biological platform to produce vitamin D, summarizing both the biological aspects of vitamin D (synthesis, ecology and evolution, metabolism, and bioequivalence) and the work done to produce it in yeast (both for vitamin D2 and for vitamin D3 ), highlighting existing challenges and potential solutions.

Genetically modified organisms: adapting regulatory frameworks for evolving genome editing technologies.

Genetic modification of living organisms has been a prosperous activity for research and development of agricultural, industrial and biomedical applications. Three decades have passed since the first genetically modified products, obtained by transgenesis, become available to the market. The regulatory frameworks across the world have not been able to keep up to date with new technologies, monitoring and safety concerns. New genome editing techniques are opening new avenues to genetic modification development and uses, putting pressure on these frameworks. Here we discuss the implications of definitions of living/genetically modified organisms, the evolving genome editing tools to obtain them and how the regulatory frameworks around the world have taken these technologies into account, with a focus on agricultural crops. Finally, we expand this review beyond commercial crops to address living modified organism uses in food industry, biomedical applications and climate change-oriented solutions.

Bispyrrolidinoindoline Epi(poly)thiodioxopiperazines (BPI-ETPs) and Simplified Mimetics: Structural Characterization, Bioactivities, and Total Synthesis.

Within the 2,5-dioxopiperazine-containing natural products generated by "head-to-tail" cyclization of peptides, those derived from tryptophan allow further structural diversification due to the rich chemical reactivity of the indole heterocycle, which can generate tetracyclic fragments of hexahydropyrrolo[2,3-b]indole or pyrrolidinoindoline skeleton fused to the 2,5-dioxopiperazine. Even more complex are the dimeric bispyrrolidinoindoline epi(poly)thiodioxopiperazines (BPI-ETPs), since they feature transannular (poly)sulfide bridges connecting C3 and C6 of their 2,5-dioxopiperazine rings. Homo- and heterodimers composed of diastereomeric epi(poly)thiodioxopiperazines increase the complexity of the family. Furthermore, putative biogenetically generated downstream metabolites with C11 and C11'-hydroxylated cores, as well as deoxygenated and/or oxidized side chain counterparts, have also been described. The isolation of these complex polycyclic tryptophan-derived alkaloids from the classical sources, their structural characterization, the description of the relevant biological activities and putative biogenetic routes, and the synthetic efforts to generate and confirm their structures and also to prepare and further evaluate structurally simple analogs will be reported.

Sparks of psychotherapeutic change: How therapists understand moments of meetings' contribution to change in psychotherapy.

OBJECTIVE: : This study describes therapists' experience of moments of meeting with their patients in psychotherapy, and how these moments relate to the patient's change process. METHOD: 13 in-depth interviews conducted with therapists from different theoretical backgrounds were analyzed using Grounded Theory. RESULTS: Four categories were established: (1) Characteristics of moments of meeting, (2) Triggers of moments of meeting (3) Enabling conditions, and (4) Main effects of moments of meeting. A conceptual model was developed around an axial phenomenon that provides understanding on how moments of meeting contribute to the change process in psychotherapy, through the construction of shared relational knowing. DISCUSSION: : The value of moments of meeting for the therapeutic process is discussed and reflected upon, as well as their effect on changes regarding implicit relational knowing.

Palladium-Catalyzed Aminocyclization-Coupling Cascades: Preparation of Dehydrotryptophan Derivatives and Computational Study.

Dehydrotryptophan derivatives have been prepared by palladium-catalyzed aminocyclization-Heck-type coupling cascades starting from o-alkynylaniline derivatives and methyl α-aminoacrylate. Aryl, alkyl (primary, secondary, and tertiary), and alkenyl substituents have been introduced at the indole C-2 position. Further variations at the indole benzene ring, as well as the C-2-unsubstituted case, have all been demonstrated. In the case of C-2 aryl substitution, the preparation of the o-alkynylaniline substrate by Sonogashira coupling and the subsequent cyclization-coupling cascade have been performed in a one-pot protocol with a single catalyst. DFT calculations have revealed significant differences in the reaction profiles of these reactions relative to those involving methyl acrylate or methacrylate, and between the reactions of the free anilines and their corresponding carbamates. Those calculations suggest that the nature of the alkene and of the acid HX released in the HX/alkene exchange step that precedes C-C bond formation could be responsible for the experimentally observed differences in reaction efficiencies.

[Ethical perspective for the COVID-19 vaccine prioritization in Chile]. / Perspectiva ética de la priorización de vacunación contra el COVID-19 en Chile.

The SARS-CoV-2 pandemic has generated an important health and economic impact on the world. Vaccines emerge as an intervention that can contribute to the control of the pandemic. Vaccines were approved for emergency use in the United States, Europe, as well as in Chile, however, they will not be immediately available, creating the need to prioritize vaccine distribution. The World Health Organization (WHO) and other international agencies established ethical frameworks to guide the distribution of the COVID-19 vaccine globally. In Chile, the Advisory Council on Vaccines and Immunizations (CAVEI) and the COVID-19 Advisory Council of the Ministry of Health (MINSAL) recommended the groups to prioritize vaccination, based on the available evidence stating that this information could change over time. In this article, we propose a reference framework of ethical principles and values to support the decision-making of prioritization and distribution of vaccines in Chile. We propose three timeless values: maximizing benefits, prioritizing the most vulnerable, reciprocity, and two transversal bioethics principles: justice and transparency. This reference framework contributes to the vaccination plan communication, the decision-making by the authorities and supports the prioritization strategy's valúes framework. With an explicit values framework we can expect better communication or priorities, a greater acceptance of SARS-CoV-2 vaccination plan by the community and an increased vaccination coverage to protect the population.

Disentangling the genetic bases of Saccharomyces cerevisiae nitrogen consumption and adaptation to low nitrogen environments in wine fermentation.

The budding yeast Saccharomyces cerevisiae has been considered for more than 20 years as a premier model organism for biological sciences, also being the main microorganism used in wide industrial applications, like alcoholic fermentation in the winemaking process. Grape juice is a challenging environment for S. cerevisiae, with nitrogen deficiencies impairing fermentation rate and yeast biomass production, causing stuck or sluggish fermentations, thus generating sizeable economic losses for wine industry. In the present review, we summarize some recent efforts in the search of causative genes that account for yeast adaptation to low nitrogen environments, specially focused in wine fermentation conditions. We start presenting a brief perspective of yeast nitrogen utilization under wine fermentative conditions, highlighting yeast preference for some nitrogen sources above others. Then, we give an outlook of S. cerevisiae genetic diversity studies, paying special attention to efforts in genome sequencing for population structure determination and presenting QTL mapping as a powerful tool for phenotype-genotype correlations. Finally, we do a recapitulation of S. cerevisiae natural diversity related to low nitrogen adaptation, specially showing how different studies have left in evidence the central role of the TORC1 signalling pathway in nitrogen utilization and positioned wild S. cerevisiae strains as a reservoir of beneficial alleles with potential industrial applications (e.g. improvement of industrial yeasts for wine production). More studies focused in disentangling the genetic bases of S. cerevisiae adaptation in wine fermentation will be key to determine the domestication effects over low nitrogen adaptation, as well as to definitely proof that wild S. cerevisiae strains have potential genetic determinants for better adaptation to low nitrogen conditions.

Indirect monitoring of TORC1 signalling pathway reveals molecular diversity among different yeast strains.

Saccharomyces cerevisiae is the main species responsible for the alcoholic fermentation in wine production. One of the main problems in this process is the deficiency of nitrogen sources in the grape must, which can lead to stuck or sluggish fermentations. Currently, yeast nitrogen consumption and metabolism are under active inquiry, with emphasis on the study of the TORC1 signalling pathway, given its central role responding to nitrogen availability and influencing growth and cell metabolism. However, the mechanism by which different nitrogen sources activates TORC1 is not completely understood. Existing methods to evaluate TORC1 activation by nitrogen sources are time-consuming, making difficult the analyses of large numbers of strains. In this work, a new indirect method for monitoring TORC1 pathway was developed on the basis of the luciferase reporter gene controlled by the promoter region of RPL26A gene, a gene known to be expressed upon TORC1 activation. The method was tested in strains representative of the clean lineages described so far in S. cerevisiae. The activation of the TORC1 pathway by a proline-to-glutamine upshift was indirectly evaluated using our system and the traditional direct methods based on immunoblot (Sch9 and Rps6 phosphorylation). Regardless of the different molecular readouts obtained with both methodologies, the general results showed a wide phenotypic variation between the representative strains analysed. Altogether, this easy-to-use assay opens the possibility to study the molecular basis for the differential TORC1 pathway activation, allowing to interrogate a larger number of strains in the context of nitrogen metabolism phenotypic differences.

Use of Raman spectroscopy in the assessment of skin after CO2 ablative fractional laser surgery on acne scars.

BACKGROUND: Ablative fractional laser surgery is a common technique for treating acne scars. However, an in vivo and noninvasive analysis of the histologic variations between acne skin and the resulting resurfaced skin is needed in order to evaluate the wound healing process of the scars induced by the ablative fractional laser surgery. MATERIALS AND METHODS: Nine patients with acne scars underwent a single treatment with a CO2 ablative fractional laser surgery. Collagen presence on the resurfaced skin was noninvasively assessed by means of Raman spectroscopy and principal component analysis. RESULTS: Principal component analysis shows that all the patients presented a collagen regeneration on the resurfaced skin after the laser treatment. CONCLUSION: Collagen plays a crucial role in the wound healing process. By assessing the collagen presence on the skin, it was possible to quantify the regenerative effects of the ablative fractional laser in a noninvasive way.

"I couldn't change the past; the answer wasn't there": A case study on the subjective construction of psychotherapeutic change of a patient with a Borderline Personality Disorder diagnosis and her therapist.

BACKGROUND: Qualitative research has provided knowledge about the subjective experiences of therapists and patients regarding the psychotherapy process and its results. Only few studies have attempted to integrate both perspectives, considering the influence of a patient's characteristics and diagnosis in the construction of this experience. AIM: To identify aspects of psychotherapy that contribute to therapeutic change based on the experience of a patient and her therapist, and to construct an integrated comprehension of the change process of a patient with Borderline Personality Disorder. METHOD: A single case was used to carry out a qualitative analysis of follow-up interviews of the participants of a long-term psychotherapy. Two qualitative approaches were combined into a model entitled "Discovery-Oriented Biographical Analysis" to reconstruct an integrated narrative. RESULTS: This method yielded an integrated narrative organized into four "chapters" that reflect the subjective construction of both the patient's and the therapist's experience of psychotherapy in terms of meaning. DISCUSSION: The understanding of psychotherapy as a multilevel process, in which different themes occur and develop simultaneously, is discussed. From this perspective, psychotherapy can be characterized as a process that involves the recovery of trust in others through corrective emotional experiences and the construction of a shared implicit relational knowledge. Clinical or methodological significance of this article: Research on the subjective experiences of psychotherapy must consider both patient and therapist as privileged but always complementary witnesses of their interaction. In addition, it should be noted that the experience of studying this biographical reconstruction generates a space where research and practice converge. The analysis of participants' narratives provides fascinating windows into their perceptions of psychotherapy and the process of change (Safran, 2013); here, the researcher is not merely a advantaged observer or a good summarizer: He/she has the chance to imbue the psychotherapy with a new meaning by connecting it with a common set of knowledge and a body of socially shared experience.

Designing Homogeneous Bromine Redox Catalysis for Selective Aliphatic C-H Bond Functionalization.

The potential of homogeneous oxidation catalysis employing bromine has remained largely unexplored. We herein show that the combination of a tetraalkylammonium bromide and meta-chloroperbenzoic acid offers a unique catalyst system for the convenient and selective oxidation of saturated C(sp3 )-H bonds upon photochemical initiation with day light. This approach enables remote, intramolecular, position-selective C-H amination as demonstrated for 20 different examples. For the first time, an N-halogenated intermediate was isolated as the active catalyst state in a catalytic Hofmann-Löffler reaction. In addition, an expeditious one-pot synthesis of N-sulfonyl oxaziridines from N-sulfonamides was developed and exemplified for 15 transformations. These pioneering examples provide a change in paradigm for molecular catalysis with bromine.

Catalytic Asymmetric Diamination of Styrenes.

An enantioselective catalytic vicinal diamination of styrenes is reported, which proceeds under entirely intermolecular reaction control. It relies on a chirally modified aryliodine(I) catalyst and proceeds within an iodine(I/III) manifold with conventional 3-chloroperbenzoic acid as a terminal oxidant. An environmentally benign solvent combination not only adds to the attractiveness of the process but also slows down the rate of the undesired background reaction. A total of 30 examples are presented, which consistently provide high enantiomeric excesses in the range 91-98%.

Dioxoiodane Compounds as Versatile Sources for Iodine(I) Chemistry.

The general synthesis, isolation and characterization of electrophilic iodine reagents of the general formula R4 N[I(O2 CAr)2 ] is reported. These compounds are air- and moisture-stable iodine(I) reagents, which were characterized including X-ray analysis. They represent conceptually new iodine(I) reagents with anions as stabilizers. These compounds display the expected performance as electrophilic reagents upon interaction with electron-rich substrates. The performance of these compounds in a total of 47 different reactions of vicinal iodooxygenation of alkenes is studied and some key features on the reagents are revealed.

[Characterization of Campylobacter jejuni samples coming form poultry meat and feces]. / Caracterización de cepas de Campylobacter jejuni obtenidas desde carne de pollo y heces de aves de corral de la zona central de Chile.

Background Campylobacter jejuni is one of the main causal agents of food borne diseases. Infections with this pathogen are mainly caused by chicken meat consumption. Aim To characterize antibiotic resistance and virulence factors in C. jejuni strains obtained from chicken meat and poultry feces in Central Chile. Material and Methods The presence of C. jejuni in 30 meat and 40 feces samples from poultry was studied. From these samples, we obtained 40 strains which were characterized at the molecular level for the presence of 16 genes involved in virulence using PCR. In parallel, antibiotic resistance for ciprofloxacin, nalidixic acid, tetracycline, erythromycin, azithromycin, chloramphenicol y ampicillin was analyzed. Results Twenty and 63% of feces and chicken meat samples were positive for C. jejuni, respectively. Moreover, a high percentage of strains showed antibiotic resistance, where 27% of strains were resistant to all tested antibiotics, except for azithromycin. Finally, 10% of the strains coming from feces contained 14 out of 16 virulence genes evaluated. Only 23% of the strains did not contain any of these genes. Conclusions A high percentage of feces and chicken meat samples are contaminated with C. jejuni. Moreover, these strains show a high genetic and phenotypic diversity represented by their antibiotic resistance profiles and the presence of virulence factors.

Disentangling the change-alliance relationship: Observational assessment of the therapeutic alliance during change and stuck episodes.

The therapeutic alliance is considered the most robust process variable associated with positive therapeutic outcome in a variety of psychotherapeutic models [Alexander, L. B., & Luborsky, L. (1986). The Penn Helping Alliance Scales. In L. S. Greenberg & W. M. Pinsoff (Eds.), The psychotherapeutic process: A research handbook (pp. 325-356). New York: Guilford Press; Horvath, A. O., Gaston, L., & Luborsky, L. (1993). The alliance as predictor of benefits of counseling and therapy. In N. Miller, L. Luborsky, J. Barber, & J. P. Docherty (Eds.), Psychodynamic treatment research: A handbook for clinical practice (pp. 247-274). New York, NY: Basic Books; Horvath, A. O., Del Re, A. C., Flückiger, C., & Symonds, D. (2011). Alliance in individual psychotherapy. Psychotherapy, 48, 9-16; Orlinky, D., Grawe, K., & Parks, B. (1994). Process and outcome in psychotherapy: Noch einmal. In A. Bergin & J. S. Garfield (Eds.), Handbook of psychotherapy and behaviour change (4th ed., pp. 270-378). New York, NY: Wiley and Sons]. The relationship between alliance and outcome has traditionally been studied based on measures that assess these therapy factors at a global level. However, the specific variations of the alliance process and their association with therapy segments that are relevant for change have not yet been fully examined. The present study examines the variations in the therapeutic alliance in 73 significant in-session events: 35 change and 38 stuck episodes identified through the observation of 14 short-term therapies of different theoretical orientations. Variations in the alliance were assessed using the VTAS-SF [Shelef, K., & Diamond, G. (2008). Short form of the revised Vanderbilt Therapeutic Alliance Scale: Development, reliability, and validity. Psychotherapy Research, 18, 433-443]. Nested analyses (HLM) indicate a statistically significant better quality of the alliance during change episodes.

Defined Palladium-Phthalimidato Catalysts for Improved Oxidative Amination.

New palladium(II)-phthalimidato complexes have been synthesized, isolated, and structurally characterized. As demonstrated from over 30 examples, they constitute superior catalysts for oxidative amination reactions of alkenes with phthalimide as the nitrogen source. This work streamlines vicinal difunctionalization of alkenes and provides access to significantly improved and experimentally simplified synthetic protocols.

Towards Uniform Iodine Catalysis: Intramolecular C-H Amination of Arenes under Visible Light.

A photochemical catalytic amination of arenes is presented. The reaction proceeds under benign iodine catalysis in the presence of visible light as the initiator and provides access to a range of differently substituted arylamines. A total of 29 examples demonstrate the broad applicability of this mild oxidation method. The scope of the reaction could further be expanded to silyl-tethered derivatives, which undergo intramolecular amination upon formation of seven-membered heterocycles. Cleavage of the silicon tether provides access to the corresponding 3-substituted anilines.

DFT Rationalization of the Diverse Outcomes of the Iodine(III)-Mediated Oxidative Amination of Alkenes.

A computational study of the mechanism for the iodine(III)-mediated oxidative amination of alkenes explains the experimentally observed substrate dependence on product distribution. Calculations with the M06 functional have been carried out on the reaction between PhI(N(SO2 Me)2 )2 and three different representative substrates: styrene, α-methylstyrene, and (E)-methylstilbene. All reactions start with electrophilic attack by a cationic PhI(N(SO2 Me)2 )(+) unit on the double bond, and formation of an intermediate with a single C-I bond and a planar sp(2) carbocationic center. The major path, leading to 1,2-diamination, proceeds through a mechanism in which the bissulfonimide initially adds to the alkene through an oxygen atom of one sulfonyl group. This behavior is now corroborated by experimental evidence. An alternative path, leading to an allylic amination product, takes place through deprotonation at an allylic C-H position in the common intermediate. The regioselectivity of this amination depends on the availability of the resonant structures of an alternate carbocationic intermediate. Only in cases where a high electronic delocalization is possible, as in (E)-methylstilbene, does the allylic amination occur without migration of the double bond.

RIM15 antagonistic pleiotropy is responsible for differences in fermentation and stress response kinetics in budding yeast.

Different natural yeast populations have faced dissimilar selective pressures due to the heterogeneous fermentation substrates available around the world; this increases the genetic and phenotypic diversity in Saccharomyces cerevisiae In this context, we expect prominent differences between isolates when exposed to a particular condition, such as wine or sake musts. To better comprehend the mechanisms underlying niche adaptation between two S. cerevisiae isolates obtained from wine and sake fermentation processes, we evaluated fermentative and fungicide resistance phenotypes and identify the molecular origin of such adaptive variation. Multiple regions were associated with fermentation rate under different nitrogen conditions and fungicide resistance, with a single QTL co-localizing in all traits. Analysis around this region identified RIM15 as the causative locus driving fungicide sensitivity, together with efficient nitrogen utilization and glycerol production in the wine strain. A null RIM15 variant confers a greater fermentation rate through the utilization of available glucose instead of its storage. However, this variant has a detrimental effect on fungicide resistance since complex sugars are not synthesized and transported into the membrane. Together, our results reveal the antagonist pleiotropic nature of a RIM15 null variant, positively affecting a series of fermentation related phenotypes, but apparently detrimental in the wild.