Un método basado en la PCR para la identificación rápida de levaduras acumuladoras de astaxantina (Phaffia spp. ) / PCR-based method for the rapid identification of astaxanthin-accumulating yeasts (Phaffia spp.)

It has been recently found that the natural distribution, habitat, and genetic diversity of astaxanthin-producing yeasts (i.e. Phaffia rhodozyma, synonym Xanthophyllomyces dendrorhous) is much greater than previously thought. P. rhodozyma is biotechnologically exploited due to its ability to produce the carotenoid pigment astaxanthin and thus, it is used as a natural source of this pigment for aquaculture. P. rhodozyma was also capable of synthesizing the potent UVB sunscreen mycosporine-glutaminol-glucoside (MGG). Therefore, further environmental studies are needed to elucidate its ecological aspects and detect new potential strains for the production of astaxanthin and MGG. However, obtaining new isolates of P. rhodozyma and related species is not always easy due to its low abundance and the presence of other sympatric and pigmented yeasts. In this work we report a successful development of a species-specific primer which has the ability to quickly and accurately detecting isolates representing all known lineages of the genus Phaffia (including novel species of the genus) and excluding closely related taxa. For this purpose, a primer of 20 nucleotides (called PhR) was designed to be used in combination with universal primers ITS3 and NL4 in a multiplex amplification. The proposed method has the sensitivity and specificity required for the precise detection of new isolates, and therefore represents an important tool for the environmental search for novel astaxanthin-producing yeasts.

Recientemente, se ha encontrado que la distribución natural, el hábitat y la diversidad genética de levaduras productoras de astaxantina (p. ej., Phaffia rhodozyma, sinónimo Xanthophyllomyces dendrorhous) son mucho mayores de lo que se pensaba. P. rhodozyma se explota biotecnológicamente debido a su capacidad para producir el pigmento carotenoide astaxantina y, por lo tanto, se utiliza como una fuente natural de este pigmento para la acuicultura. También se encontró que esta levadura es capaz de sintetizar el potente protector solar UVB micosporina-glutaminol-glucósido (MGG). Por lo tanto, más estudios ambientales para dilucidar sus aspectos ecológicos y detectar nuevas cepas potenciales productoras de astaxantina y MGG son necesarios. Sin embargo, la obtención de nuevos aislamientos de P. rhodozyma y especies relacionadas no siempre es fácil debido a su baja abundancia y a la presencia de otras levaduras simpátricas y pigmentadas. En este trabajo se describe el desarrollo exitoso de un cebador especie-específico que tiene la capacidad de detectar rápidamente y con precisión cepas representativas de todos los linajes del género Phaffia previamente reportados (incluyendo nuevas especies del género) y excluir especies estrechamente relacionadas. Para ello, se diseñó un cebador de 20 nucleótidos (denominado PhR) para ser utilizado en combinación con los cebadores universales ITS3 y NL4 en una amplificación multiplex. El método propuesto tiene la sensibilidad y la especificidad requerida para la detección precisa de nuevos aislamientos y, por lo tanto, representa una importante herramienta para la búsqueda ambiental de nuevas levaduras productoras de astaxantina.

The diversity, extracellular enzymatic activities and photoprotective compounds of yeasts isolated in Antarctica

The diversity of yeasts collected from different sites in Antarctica (Admiralty Bay, King George Island and Port Foster Bay and Deception Island) and their ability to produce extracellular enzymes and mycosporines were studied. Samples were collected during the austral summer season, between November 2006 and January 2007, from the rhizosphere of Deschampsia antarctica, ornithogenic (penguin guano) soil, soil, marine and lake sediments, marine water and freshwater from lakes. A total of 89 isolates belonging to the following genera were recovered: Bensingtonia, Candida, Cryptococcus, Debaryomyces, Dioszegia, Exophiala, Filobasidium, Issatchenkia (Pichia), Kodamaea, Leucosporidium, Leucosporidiella, Metschnikowia, Nadsonia, Pichia, Rhodotorula, and Sporidiobolus, and the yeast-like fungi Aureobasidium, Leuconeurospora and Microglossum. Cryptococcus victoriae was the most frequently identified species. Several species isolated in our study have been previously reported to be Antarctic psychophilic yeasts, including Cr. antarcticus, Cr. victoriae, Dioszegia hungarica and Leucosporidium scottii. The cosmopolitan yeast species A. pullulans, C. zeylanoides, D. hansenii, I. orientalis, K. ohmeri, P. guilliermondii, Rh. mucilaginosa, and S. salmonicolor were also isolated. Five possible new species were identified. Sixty percent of the yeasts had at least one detectable extracellular enzymatic activity. Cryptococcus antarcticus, D. aurantiaca, D. crocea, D. hungarica, Dioszegia sp., E. xenobiotica, Rh. glaciales, Rh. laryngis, Microglossum sp. 1 and Microglossum sp. 2 produced mycosporines. Of the yeast isolates, 41.7 percent produced pigments and/or mycosporines and could be considered adapted to survive in Antarctica. Most of the yeasts had extracellular enzymatic activities at 4ºC and 20ºC, indicating that they could be metabolically active in the sampled substrates.