De novo production of six key grape aroma monoterpenes by a geraniol synthase-engineered S. cerevisiae wine strain

[Background] Monoterpenes are important contributors to grape and wine aroma. Moreover, certain monoterpenes have been shown to display health benefits with antimicrobial, anti-inflammatory, anticancer or hypotensive properties amongst others. The aim of this study was to construct self-aromatizing wine yeasts to overproduce de novo these plant metabolites in wines.[Results] Expression of the Ocimum basilicum (sweet basil) geraniol synthase (GES) gene in a Saccharomyces cerevisiae wine strain substantially changed the terpene profile of wine produced from a non-aromatic grape variety. Under microvinification conditions, and without compromising other fermentative traits, the recombinant yeast excreted geraniol de novo at an amount (~750 μg/L) well exceeding (>10-fold) its threshold for olfactory perception and also exceeding the quantities present in wines obtained from highly aromatic Muscat grapes. Interestingly, geraniol was further metabolized by yeast enzymes to additional monoterpenes and esters: citronellol, linalool, nerol, citronellyl acetate and geranyl acetate, resulting in a total monoterpene concentration (~1,558 μg/L) 230-fold greater than that of the control. We also found that monoterpene profiles of wines derived from mixed fermentations were found to be determined by the composition of the initial yeast inocula suggesting the feasibility of producing ‘à la carte’ wines having predetermined monoterpene contents.[Conclusions] Geraniol synthase-engineered yeasts demonstrate potential in the development of monoterpene enhanced wines.This work was funded by the Spanish MINECO/FEDER Grants CSD2007-0063 and AGL2011-29925. We also acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI). E. Pardo was financially supported by JCI2007-123/733 and JAEDOC086-FSE contracts. J. Rico was the recipient of a FPI (Formación de Personal de Investigador) pre-doctoral fellowship from the Comisión Interministerial de Ciencia y Tecnología (CICYT).Peer Reviewe

Urticaria and silent parasitism by Ascaridoidea: Component-resolved diagnosis reinforces the significance of this association

Urticaria remains a major problem in terms of aetiology, investigation, and management, and although parasitic diseases are considered potential causes, the absence of a consistent link between parasitic infections and skin allergy symptoms leads to the need for a deeper study of parameters that support this association. The objectives of this study were to analyse a possible relationship between parasitism by Ascarididae (Toxocara canis and Anisakis simplex) and the clinical expression of urticaria and to identify possible parasitic molecular markers for improving the diagnosis of unknown urticaria aetiology. The prevalence of Toxocara and Anisakis infestations was evaluated by measuring the levels of specific IgG (sIgG) and IgE (sIgE) antibodies against crude extracts and isolated components from whole larvae of Anisakis simplex (Ani s 1, Ani s 3 and Ani s 7) and Toxocara canis (TES-120, TES-70, TES-32 and TES-26) using immunologic and molecular diagnostic methods. A cross-sectional study was performed in a group of 400 individuals. The study group consisted of 95 patients diagnosed with urticaria (55 with chronic urticaria and 40 with acute urticaria). A control group consisted of 305 subjects without urticaria (182 diagnosed with respiratory allergy and 123 without allergy). Statistically significant differences were demonstrated in the seroprevalence of specific IgG and IgE antibodies between the urticaria patients and the healthy general population when isolated ascarid antigens were evaluated. The prevalence of IgG antibodies against Ani s 1, IgE antibodies against TES-120 and IgE antibodies against TES-70 were significantly different between the control individuals (healthy general population) and patients with urticaria. Moreover, the urticaria patient group demonstrated a higher seroprevalence of antibodies (sIgE and sIgG) against Anisakis simplex larva whole extract than the control group but just with statistically diferences when sIgE was evaluated. The presence of IgE and/or IgG antibodies against Ani s 3 (tropomyosin) can help to discriminate between patients with and without urticaria. Both ascarids seem to be associated with urticaria, although in our region, Anisakis seems to have greater involvement than Toxocara in this relationship. Molecular diagnostics can be used to associate urticaria with parasite infestations. Tropomyosin and Ani s 1 were the most relevant markers to demonstrate the association between urticaria and the most relevant Ascarididae parasites in our region. Author summary Urticaria remains a major problem in terms of aetiology, investigation, and management. It seems that some parasitic diseases could be causes of urticaria. Although there is no definitive consistent link between both entities, several studies have shown a significant positive association between parasite infections, mainly helminthic infections, and urticaria. The study of prevalence data for the most relevant Ascarididae infections in our region in different groups (healthy general population, respiratory allergy patients and patients with urticaria), through the measure of specific IgE and IgG antibodies against crude and molecular antigens of Anisakis and Toxocara, allowed us to define some antigenic markers that can be used to discriminate urticaria patients from the control individuals.This research was supported by funds to support the Activities of the Research Groups of the Basque University System (Project IT-1043-16), Department of Education, Universities and Research, Government of The Basque Country Spain. The funders had no role in study design, data collection and analysis, decision to publish, or preparation manuscript

L-Rhamnose induction of Aspergillus nidulans α-L-rhamnosidase genes is glucose repressed via a CreA-independent mechanism acting at the level of inducer uptake

<p>Abstract</p> <p>Background</p> <p>Little is known about the structure and regulation of fungal α-L-rhamnosidase genes despite increasing interest in the biotechnological potential of the enzymes that they encode. Whilst the paradigmatic filamentous fungus <it>Aspergillus nidulans </it>growing on L-rhamnose produces an α-L-rhamnosidase suitable for oenological applications, at least eight genes encoding putative α-L-rhamnosidases have been found in its genome. In the current work we have identified the gene (<it>rhaE</it>) encoding the former activity, and characterization of its expression has revealed a novel regulatory mechanism. A shared pattern of expression has also been observed for a second α-L-rhamnosidase gene, (AN10277/<it>rhaA</it>).</p> <p>Results</p> <p>Amino acid sequence data for the oenological α-L-rhamnosidase were determined using MALDI-TOF mass spectrometry and correspond to the amino acid sequence deduced from AN7151 (<it>rhaE</it>). The cDNA of <it>rhaE </it>was expressed in <it>Saccharomyces cerevisiae </it>and yielded <it>p</it>NP-rhamnohydrolase activity. Phylogenetic analysis has revealed this eukaryotic α-L-rhamnosidase to be the first such enzyme found to be more closely related to bacterial rhamnosidases than other α-L-rhamnosidases of fungal origin. Northern analyses of diverse <it>A. nidulans </it>strains cultivated under different growth conditions indicate that <it>rhaA </it>and <it>rhaE </it>are induced by L-rhamnose and repressed by D-glucose as well as other carbon sources, some of which are considered to be non-repressive growth substrates. Interestingly, the transcriptional repression is independent of the wide domain carbon catabolite repressor CreA. Gene induction and glucose repression of these <it>rha </it>genes correlate with the uptake, or lack of it, of the inducing carbon source L-rhamnose, suggesting a prominent role for inducer exclusion in repression.</p> <p>Conclusions</p> <p>The <it>A. nidulans rhaE </it>gene encodes an α-L-rhamnosidase phylogenetically distant to those described in filamentous fungi, and its expression is regulated by a novel CreA-independent mechanism. The identification of <it>rhaE </it>and the characterization of its regulation will facilitate the design of strategies to overproduce the encoded enzyme - or homologs from other fungi - for industrial applications. Moreover, <it>A. nidulans </it>α-L-rhamnosidase encoding genes could serve as prototypes for fungal genes coding for plant cell wall degrading enzymes regulated by a novel mechanism of CCR.</p

Fungal Allergen and Mold Allergy Diagnosis: Role and Relevance of Alternaria alternata Alt a 1 Protein Family

Alternaria is a genus of worldwide fungi found in different habitats such as soil, the atmosphere, plants or indoor environments. Alternaria species are saprobic—largely involved in the decomposition of organic material—but they can also act as animal pathogens, causing disease in humans and animals, developing infections, toxicosis and allergic diseases. A. alternata is considered one of the most important sources of fungal allergens worldwide and it is associated with severe asthma and respiratory status. Among the A. alternata allergens, Alt a 1 is the main sensitizing allergen and its usefulness in diagnosis and immunotherapy has been demonstrated. Alt a 1 seems to define a protein family that can be used to identify related pathogenic fungi in plants and fruits, and to establish taxonomic relationships between the different fungal divisions.This work was supported by the Basque Country Government: Consolidated Research Groups of the Basque University Research System: Project IT-1043-16 and the Health Research Department of the Basque Government: Projects: SAN18/16 and SAN20/04

Purified Native and Recombinant Major Alternaria alternata Allergen (Alt a 1) Induces Allergic Asthma in the Murine Model

Aeroallergens such us the spores of Alternaria alternata are described as the most important agents associated with respiratory allergies and severe asthma. Various experimental models of asthma have been developed using A. alternata extracts to study the pathogenesis of asthma, establishing the main parameters that trigger the asthmatic response. In this study, we describe a mouse model of asthma induced only by Alt a 1. To induce the allergic response, mice were challenged intranasally with the major allergen of A. alternata, Alt a 1. The presence of eosinophils in the lungs, elevated concentrations of Th2 family cytokines, lymphocyte proliferation and elevated IgE total serum levels indicated that the sensitisation and challenge with Alt a 1 induced the development of airway inflammation. Histological studies showed an eosinophilic cellular infiltrate in the lung tissue of mice instilled with Alt a 1. We demonstrate that Alt a 1 alone is capable of inducing a lung inflammatory response with an increase in IgE serum levels mimicking the allergic asthma immunoresponse when it is administered into BALB/c mice. This model will allow the evaluation of the immunoregulatory or immunotolerant capacity of several molecules that can be used in targeted immunotherapy for fungal allergic asthmaThis work was supported by the Government of the Basque Country, Department of Research and Education, Linguistic Policy and Culture in the Program: Grants for University Research Groups (Project IT-1043-16) and by the Department of Health (Projects SAN18/16 and SAN20/04)

Two-stage case-control association study of dopamine-related genes and migraine

Background We previously reported risk haplotypes for two genes related with serotonin and dopamine metabolism: MAOA in migraine without aura and DDC in migraine with aura. Herein we investigate the contribution to migraine susceptibility of eight additional genes involved in dopamine neurotransmission. Methods We performed a two-stage case-control association study of 50 tag single nucleotide polymorphisms (SNPs), selected according to genetic coverage parameters. The first analysis consisted of 263 patients and 274 controls and the replication study was composed by 259 cases and 287 controls. All cases were diagnosed according to ICHD-II criteria, were Spanish Caucasian, and were sex-matched with control subjects. Results Single-marker analysis of the first population identified nominal associations of five genes with migraine. After applying a false discovery rate correction of 10%, the differences remained significant only for DRD2 (rs2283265) and TH (rs2070762). Multiple-marker analysis identified a five-marker T-C-G-C-G (rs12363125-rs2283265-rs2242592-rs1554929-rs2234689) risk haplotype in DRD2 and a two-marker A-C (rs6356-rs2070762) risk haplotype in TH that remained significant after correction by permutations. These results, however, were not replicated in the second independent cohort. Conclusion The present study does not support the involvement of the DRD1, DRD2, DRD3, DRD5, DBH, COMT, SLC6A3 and TH genes in the genetic predisposition to migraine in the Spanish population

The Aspergillus niger major allergen (Asp n 3) DNA-specific sequence is a reliable marker to identify early fungal Contamination and postharvest damage in Mangifera indica fruit

[EN] The aim of this work was to study the value of the main allergen Asp n 3 of Aspergillus niger as a molecular marker of allergenicity and pathogenicity with the potential to be used in the identification of A. niger as a contaminant and cause of spoilage of Mangifera indica. Real-time polymerase chain reaction (RT-PCR) was used for the amplification of Asp n 3 gene. Two pairs of primers were designed: one for the amplification of the entire sequence and another one for the amplification of the most conserved region of this peroxisomal protein. The presence of A. niger was demonstrated by the early detection of the allergenic protein Asp n 3 coding gene, which could be considered a species-specific marker. The use of primers designed based on the conserved region of the Asp n 3 encoding gene allowed us to identify the presence of the closely related fungal species Aspergillus fumigatus by detecting Asp n 3 homologous protein, which can be cross-reactive. The use of conserved segments of the Asp n 3 gene or its entire sequence allows us to detect phylogenetically closely related species within the Aspergilaceae family or to identify species-specific contaminating fungi.This work was supported by the Government of the Basque Country, Department of Research and Education, Linguistic Policy and Culture in the Program: Grants for University Research Groups (project IT-1043-16)

Identification of the main allergen sensitizers in an Iran asthmatic population by molecular diagnosis

Background: There has been a significant growth in the prevalence of allergy, mainly associated to IgE-mediated disorders such as asthma and rhinitis. The identification of atopy in asthmatic patients through the measurement of specific IgE can help to identify risk factors that cause asthmatic symptoms in patients. The development and use of individualized allergen-based tests by the Component Resolved Diagnosis has been a crucial advance in the accurate diagnosis and control of allergic patients. The objective of this work was to assess the usefulness of molecular diagnosis to identify environmental allergens as possible factors influencing the development and manifestation of asthma in a group of asthmatic patients from Iran. Methods: Studied population: 202 adult asthmatic patients treated at the Loghman Hakim Hospital and Pasteur Institute of Teheran (Iran) from 2011 to 2012. Specific IgE determined by the ImmunoCAP system were used to both evaluate the patients' atopic condition and the molecules involved in the allergic sensitization. SDS-PAGE IgE-immunoblotting associated with mass spectrometry was carried out to study the cockroach IgE-binding sensitizing proteins. Results: Forty-five percent of all patients could be considered atopic individuals. Eighty-two percent of atopic patients were sensitized to pollen allergens. The Salsola kali (Sal k 1) and the Phleum pratense (rPhl p 1 and/or rPhl p 5) major allergens were the most common sensitizers among pollens (71% and 18%, respectively). Thirty-five percent of the atopic population was sensitized to cockroach. Four different allergens, including a previously unknown alpha-amylase, were identified in the cockroach extract. No significant associations could be demonstrated between the severity of asthma and the specific IgE levels in the atopic population. Statistical analysis identified the Sal k 1 as the main protein allergen influencing the development and expression of asthma in the studied population. Conclusions: Pollen and cockroach were the most relevant allergen sources in the asthmatic population. The Salsola kali major allergen was the main cause for sensitization in the atopic patients suffering asthma. Using the Component Resolved Diagnosis, it was possible to identify a new Blattella germanica cockroach allergen (Blattella alpha amylase 53 kDa) that could sensitize a relevant percentage of this population.This study has been founded by the Government of The Basque Country, Project IT787-13

Interiorización del método científico en alimentación sostenible mediante la experimentación en el aula

[ES] La alimentación sostenible hace referencia a aquella alimentación saludable que se adapta al entorno y la cultura, que disminuye el impacto ambiental, respeta los recursos naturales y la biodiversidad y es económicamente accesible. El alumnado que cursa alguno de los grados de Ciencias de la Alimentación debe ser consciente del concepto de sostenibilidad, integrarlo en su vida diaria, así como saber transmitirlo a la sociedad. Este alumnado busca adquirir conocimientos a través de metodologías diferentes y complementarias. La combinación de prácticas tradicionales y modernas son vitales para promover diferentes habilidades cognitivas en el estudiantado. Es importante aumentar su conocimiento y sus competencias en la resolución de problemas pero también lo es incrementar sus habilidades de razonamiento. Teniendo en cuenta que los y las estudiantes en Ciencias de la Alimentación deben trabajar desde la aplicación del método científico, una de las maneras más adecuadas de interiorizar este es llevándolo a la práctica. Por ello, en este proyecto se pretende que el alumnado sea capaz de plantear una hipótesis de trabajo en alimentación sostenible en consonancia con los ODS relacionados y llevar a cabo un taller práctico que potencie sus competencias comunicativas en el ámbito universitario.[EN] Sustainable food refers to healthy food that is adapted to the environment and culture, that reduces environmental impact, respects natural resources and biodiversity and is economically accessible. Students taking a degree in Food Science must be aware of the concept of sustainability, integrate it into their daily lives and know how to transmit it to society. These students want to acquire knowledge through different and complementary methodologies. The combination of traditional and modern practices are vital to promote different cognitive skills in students. It is important to increase their knowledge and problem solving competences but it is also important to increase their reasoning skills. Taking into account that students in Food Science must work from the application of the scientific method, one of the most appropriate ways to learn this method is by putting it into practice. Therefore, the aim of this project is to ensure that students are able to propose a working hypothesis on sustainable food in line with the related SDGs and to carry out a practical workshop that enhances their communicative skills in the university environment.Gandía Gómez, M.; Rodríguez-Carrasco, Y.; Cabrera-Pastor, A.; Pardo, E.; Gamero, A. (2023). Interiorización del método científico en alimentación sostenible mediante la experimentación en el aula. Editorial Universitat Politècnica de València. 581-589. https://doi.org/10.4995/INRED2023.2023.1653758158