Luis B. Flores-Cotera

Diversity of endophytic fungi of Taxus globosa (Mexican yew)

The aim of this work was the isolation and taxonomic characterization of endophytic fungi from Taxus globosa at the Sierra Alta Hidalguense, Mexico. A total of 116 fungi were isolated from the bark, branches, leaves and roots of healthy yew trees. Based on morphological characteristics 57 were selected for taxonomic characterization through phylogenetic analysis of their 28S rDNA sequences. The fungal isolates belonged to Ascomycota (77.2%) and Basidiomycota (22.8%). Twelve different fungal groups were identified: Coniochaetales, Eurotiales, Hypocreales, Phyllachorales, Pleosporales, Pezizales, Sordariomycetidae, Sordariales, Trichosphaeriales, Xylariales, Agaricales and Polyporales. The taxa Alternaria sp. Aspergillus sp., Cochliobolus sp., Coprinellus domesticus, Hypoxylon sp., Polyporus arcularius, Xylaria juruensis and Xylariaceae were the most frequently isolated. The genera Annulohypoxylon, Cercophora, Conoplea, Daldinia, Lecythophora, Letendraea, Massarina, Phialophorophoma, Sporormia, Xylomelasma, Coprinellus, Polyporus and Trametes for the first time were isolated from yews; this suggests that T. globosa harbours novel and highly diverse fungi. The Shannon-Weaver and Simpson diversity index values for the overall fungal community were H′ = 3.139 and 1-D = 0.941 respectively.

Microbial paclitaxel: advances and perspectives

Paclitaxel is a potent and widely used antitumor agent. Considerable worldwide research efforts have been carried out on different production alternatives. Since the description of the first paclitaxel-producing fungi, more than 15 years ago, microorganisms have been investigated as potential alternatives for an environmentally acceptable, relatively simple and inexpensive method to produce paclitaxel. However, in spite of significant research on paclitaxel-producing microorganisms, no commercial fermentation process has been implemented up to now. The aim of this study is to review the present status of research on paclitaxel-producing microorganisms and the ongoing efforts to develop heterologous paclitaxel biosynthesis, and analyze the perspectives of microbial fermentation for paclitaxel production.

Molecular phylogeny and paclitaxel screening of fungal endophytes from Taxus globosa.

We studied the endophytic mycoflora associated with Taxus globosa, the Mexican yew. The study localities; Las Avispas (LA), San Gaspar (SG), and La Mina (LM) were three segments of cloud forest within the range of Sierra Gorda Biosphere Reserve, México. Overall, 245 endophytes were isolated and 105 representative Ascomycota (morphotaxons) were chosen for phylogenetic and genotypic characterization. Maximum likelihood analyses of large subunit of ribosomal RNA (LSU) rDNA showed well-supported clades of Dothideomycetes, Eurotiomycetes, Leotiomycetes, Pezizomycetes, and Sordariomycetes. Analyses of ITS rDNA groups showed 57 genotypes (95% sequence similarity), in general consistent with the phylogenetically delimitated taxa based on LSU rDNA sequences. The endophyte diversity measured by Fishers α, Shanonn, and Simpson indices was ca. three-fold and ca. two-fold greater in LM than in LA and SG respectively. A screening for paclitaxel using a competitive inhibition enzyme immunoassay showed 16 positive isolates producing between 65 and 250 ng l(-1). The isolates included Acremonium, Botryosphaeria, Fusarium, Gyromitra, Nigrospora, Penicillium, three novel Pleosporales, and Xylaria.

Continuous Cr(VI) removal by Scenedesmus incrassatulus in an airlift photobioreactor

Cr(VI) removal by Scenedesmus incrassatulus was characterized in a continuous culture system using a split-cylinder internal-loop airlift photobioreactor fed continuously with a synthetic effluent containing 1.0mg Cr(VI) l(-1) at dilution rate (D) of 0.3d (-1). At steady state, there was a small increase (6%) on the dry biomass (DB) concentration of Cr(VI)-treated cultures compared with the control culture. 1.0mg Cr(VI) l(-1) reduced the photosynthetic pigments content and altered the cellular morphology, the gain in dry weight was not affected. At steady state, Cr(VI) removal efficiency was 43.5+/-1.0% and Cr(VI) uptake was 1.7+/-0.1 mg Cr(VI) g(-1) DB. The system reached a specific metal removal rate of 458 microg Cr(VI) g(-1) DB d(-1), and a volumetric removal rate of 132 microg Cr(VI) l(-1) d(-1).

ATP-citrate lyase activity and carotenoid production in batch cultures of Phaffia rhodozyma under nitrogen-limited and nonlimited conditions

ATP-citrate lyase (ACL) is the key cytoplasmic enzyme which supplies acetyl-CoA for fatty acids in oleaginous yeast. Although it has been suggested that fatty acid and carotenoid biosynthesis may have a common source of acetyl-CoA in Phaffia rhodozyma, the source for carotenoids is currently unknown. The purpose of this work was to analyze the development of ACL activity during batch cultures of P. rhodozyma under ammonium-limited and nonammonium-limited conditions and study its possible relationship with carotenoid synthesis. Every experiment showed carotenoid accumulation linked to an increasing ACL activity. Moreover, the ACL activity increased with dissolved oxygen (DO), i.e., ACL responded to DO in a similar way as carotenoid synthesis. Additionally, in the ammonium-limited culture, ACL activity increased upon ammonium depletion. However, the contribution to carotenoid accumulation in that case was negligible. This suggests that P. rhodozyma has developed two components of ACL, each one responsive to a different environmental stimulus, i.e., DO and ammonium depletion. The role of each component is still unknown; however, considering that the former responds to DO and the known role of carotenoids as antioxidants, it may be a provider of acetyl-CoA for carotenoid synthesis.

A possible mechanism of metabolic regulation in Gibberella fujikuroi using a mixed carbon source of glucose and corn oil inferred from analysis of the kinetics data obtained in a stirrer tank bioreactor

A nonstructured model was used to study the dynamics of gibberellic acid production in a stirred tank bioreactor. Experimental data were obtained from submerged batch cultures of Gibberella fujikuroi (CDBB H‐984) grown in varying ratios of glucose‐corn oil as the carbon source. The nitrogen depletion effect was included in mathematical model by considering the specific kinetic constants as a linear function of the normalized nitrogen consumption rate. The kinetics of biomass growth and consumption of phosphate and nitrogen were based on the logistic model. The traditional first‐order kinetic model was used to describe the specific consumption of glucose and corn oil. The nitrogen effect was solely included in the phosphate and corn oil consumption and biomass growth. The model fit was satisfactory, revealing the dependence of the kinetics with respect to the nitrogen assimilation rate. Through simulations, it was possible to make diagrams of specific growth rate and specific rate of substrate consumptions, which was a powerful tool for understanding the metabolic interactions that occurred during the various stages of fermentation process. This kinetic analysis provided the proposal of a possible mechanism of regulation on growth, substrate consumptions, and production of gibberellic acid (GA3) in G. fujikuroi.

A common mechanism explains the induction of aerobic fermentation and adaptive antioxidant response in Phaffia rhodozyma

BackgroundGrowth conditions that bring about stress on Phaffia rhodozyma cells encourage the synthesis of astaxanthin, an antioxidant carotenoid, which protects cells against oxidative damage. Using P. rhodozyma cultures performed with and without copper limitation, we examined the kinetics of astaxanthin synthesis along with the expression of asy, the key astaxanthin synthesis gene, as well as aox, which encodes an alternative oxidase protein.ResultsCopper deficiency had a detrimental effect on the rates of oxygen consumption and ethanol reassimilation at the diauxic shift. In contrast, copper deficiency prompted alcoholic fermentation under aerobic conditions and had a favorable effect on the astaxanthin content of cells, as well as on aox expression. Both cultures exhibited strong aox expression while consuming ethanol, but particularly when copper was absent.ConclusionWe show that the induction of either astaxanthin production, aox expression, or aerobic fermentation exemplifies the crucial role that redox imbalance plays in triggering any of these phenomena. Based on our own results and data from others, we propose a mechanism that rationalizes the central role played by changes of respiratory activity, which lead to redox imbalances, in triggering both the short-term antioxidant response as well as fermentation in yeasts and other cell types.