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The yeast Yarrowia lipolytica is distantly related to Saccharomyces cerevisiae, can be genetically modified, and can grow in both haploid and diploid states in either yeast, pseudomycelial, or mycelial forms, depending on environmental conditions. Previous results have indicated that the STE and RIM pathways, which mediate cellular switching in other dimorphic yeasts, are not required for Y. lipolytica morphogenesis. To identify the pathways involved in morphogenesis, we mutagenized a wild-type strain of Y. lipolytica with a Tn3 derivative. We isolated eight tagged mutants, entirely defective in hyphal formation, from a total of 40,000 mutants and identified seven genes homologous to S. cerevisiae CDC25, RAS2, BUD6, KEX2, GPI7, SNF5, and PPH21. We analyzed their abilities to invade agar and to form pseudomycelium or hyphae under inducing conditions and their sensitivity to temperature and to Calcofluor white. Chitin staining was used to detect defects in their cell walls. Our results indicate that a functional Ras-cyclic AMP pathway is required for the formation of hyphae in Y. lipolytica and that perturbations in the processing of extracellular, possibly parietal, proteins result in morphogenetic defects.

The biotechnological potential of Yarrowia lipolytica, as a single cell oil-producing microorganism, is presented in this review. Although initially this yeast species was considered as a lipid-degrading, recently, it was reclassified as a lipid-producing microorganism, since it has been reported to be capable of accumulating diverse desirable fatty acids after metabolic pathway engineering. In the first part of the present document, a general revision of the oil metabolic pathways and the capacity of oil production in Y. lipolytica is presented. The single cell oil produced by these metabolic engineering strategies has been designed by optimization, introduction, or suppression of new pathways to increase yield on lipid production. Later on, the genetic regulation systems and the lipid composition generated by this yeast for industrial purposes are discussed. These lipids could be safely used in the chemical food and biofuel industries, due to their high proportion of oleic acid. This document emphasizes in the overviewing at Y. lipolytica as an ideal oil cell factory, and as an excellent model to produce single cell oil.

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Chitin is the second most abundant organic compound in nature and represents a rich carbon and nitrogen source that is primarily transformed by bacterial communities. Bacteria capable of gradually hydrolyzing chitin into N-acetylglucosamine monomers can have applications in the transformation of residues from shrimp and other crustaceans. The objective of the present study was to isolate, characterize and identify microorganisms with high chitinolytic activity. These microorganisms were isolated and characterized based on macro- and microscopic morphological traits. Strains were selected on colloidal chitin agar medium primarily based on a hydrolysis halo larger than 2 mm and a growing phase no longer than 6 days. Secondary selection consisted of semi-quantitative evaluation of chitinolytic activity with a drop dilution assay. From the above, ten strains were selected. Then, strain-specific activity was evaluated. The B4 strain showed the highest specific activity, which was 6,677.07 U/mg protein. Molecular identification indicated that the isolated strains belong to the species Stenotrophomonas maltophilia.

White spot syndrome virus (WSSV) is the most aggressive disease affecting cultured shrimp. One possibility to tackle it is by means of RNA interference (RNAi) induced by the presence of double-stranded RNA (dsRNA). Normally, dsRNA is a product of the cellular machinery to gene regulation, but it can be produced synthetically and introduced into specific tissues or cells and thereby induce RNAi. Although in vitro production of dsRNA is possible, this is high cost. An alternative is to produce dsRNA in vivo using biological systems such as bacteria or yeasts. In this regard, Yarrowia lipolytica offers distinctive advantages for dsRNA production. The objective was to develop a Y. lipolytica strain able to produce dsRNA-specific against WSSV and to evaluate its antiviral activity in the white leg shrimp Litopenaeus vannamei.From the 0.4 and 0.6 Kb fragments of the ORF89 gene, a dsRNA-ORF89-producing construct was built in the plasmid pJC410; the resulting construct (pARY410) was used to transform Y. lipolytica to drive the specific expression of dsRNA-ORF89.Yeast colonies positive to the WSSV-ORF89 gene were selected. The expression of dsRNA-ORF89 and RNAse III was measured being detected at 32 and 48 hr. Subsequently, the antiviral activity of dsRNA-ORF89 was tested in a WSSV challenge bioassay. The results showed survival in dsRNA-ORF89 shrimp (25%) compared to control organisms treated with total RNA from the yeast P01-AS harvested at 32 hr. In conclusion, Y. lipolytica is a convenient host to produce and deliver dsRNA-ORF89 able to protect WSSV-challenged shrimp. K E Y W O R D Santiviral, Litopenaeus vannamei, ORF89, RNAi, RNAse III

Composite films with Aloe vera (A), chitosan (Ch) and essential oils (EOs) were formulated. Six of the twelve combinations tested formed films: A70Ch30, A70Ch30-15, A60Ch40, A60Ch40-15, A50Ch50, and A50Ch50-15. The A60Ch40-15 film showed the best physicochemical characteristics as well as the greatest in vitro antifungal activity. Although the A90Ch10 and A80Ch20-15 mixtures did not form films, their solutions showed high antifungal activity in vitro. Based on multivariate analysis of the data, A60Ch40-15, A90Ch10 and A80Ch20-15 films were selected as coating treatments for papaya during storage at 30 ± 2 °C and 80% RH. Uncoated fruits (control 1) and treated with synthetic fungicide (control 2) were used as control. Coated fruits showed lower respiration rate, greater firmness and fewer changes in external coloration compared to control. Furthermore, these coatings reduced the incidence and severity of fungal disease by 40-50% compared to control 2. Aloe vera-chitosan films (A90Ch10 and A60Ch40-15), enriched with the EOs of cinnamon (10 mL L -1 ) and thyme (10 mL L -1 ), improved quality of the fruit (higher firmness, lower CO 2 content, less internal color change) with 50% less disease incidence during storage at room temperature.