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BackgroundBacteria may compete with yeast for nutrients during bioethanol production process, potentially causing economic losses. This is the first study aiming at the quantification and identification of Lactic Acid Bacteria (LAB) present in the bioethanol industrial processes in different distilleries of Brazil.ResultsA total of 489 LAB isolates were obtained from four distilleries in 2007 and 2008. The abundance of LAB in the fermentation tanks varied between 6.0 × 105 and 8.9 × 108 CFUs/mL. Crude sugar cane juice contained 7.4 × 107 to 6.0 × 108 LAB CFUs. Most of the LAB isolates belonged to the genus Lactobacillus according to rRNA operon enzyme restriction profiles. A variety of Lactobacillus species occurred throughout the bioethanol process, but the most frequently found species towards the end of the harvest season were L. fermentum and L. vini. The different rep-PCR patterns indicate the co-occurrence of distinct populations of the species L. fermentum and L. vini, suggesting a great intraspecific diversity. Representative isolates of both species had the ability to grow in medium containing up to 10% ethanol, suggesting selection of ethanol tolerant bacteria throughout the process.ConclusionsThis study served as a first survey of the LAB diversity in the bioethanol process in Brazil. The abundance and diversity of LAB suggest that they have a significant impact in the bioethanol process.

The present work aims at characterizing the regulatory mechanisms of metabolism and product formation of BHK cells producing a recombinant antibody/cytokine fusion protein. This work was carried out through the achievement of several steady‐states in chemostat cultures, corresponding to different glucose and glutamine levels in the feed culture medium. Results obtained indicate that both glucose and glutamine consumptions show a Michaelis–Menten dependence on residual glucose and glutamine concentrations, respectively. Similar dependence was also observed for lactate and ammonia productions. K GlcGlc and K GlnGln were estimated to be 0.4 and 0.15 mM, respectively, while q maxGlc and q maxGln were estimated to be 1.8 and 0.55 nmol 10−6cells min−1, respectively. At very low glucose concentrations, the glucose‐to‐lactate yield decreased markedly showing a metabolic shift towards lower lactate production; also, the glucose‐to‐cells yield was increased. At very low‐glutamine concentrations, the glutamine‐to‐ammonia and glutamine‐to‐cells yields increased, showing a more efficient glutamine metabolism. Overall, amino acid consumption was increased under low glucose or glutamine concentrations. Metabolic‐flux analysis confirmed the metabolic shifts by showing increases in the fluxes of the more energetically efficient pathways, at low‐nutrient concentrations. No effect of glucose or glutamine concentrations on the cell‐specific productivity was observed, even under metabolically shifted metabolism; therefore, it is possible to confine the cells to a more efficient metabolic state maintaining the productivity of the recombinant product of interest, and consequently, increasing final product titers by increasing cell concentration and culture length. This work is intended to be a model approach to characterize cell metabolism in an integrated way; it is highly valuable for the establishment of operating strategies in mammalian cell fermentations in which cell metabolism is to be confined to a desired state. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 66: 104–113, 1999.

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In this work, a BHK21 clone producing a recombinant antibody/cytokine fusion protein was used to study the dependence of cell metabolism on the glucose and glutamine levels in the culture medium. Results obtained indicate that both glucose and glutamine consumptions show a Michaelis-Menten dependence on glucose and glutamine concentrations respectively. A similar dependence is also observed for lactate and ammonia productions. The estimated value of the Michaelis constant for the dependence of lactate production on glucose (KLacGlc) was 1.4 +/- 0.1 mM and for the dependence of ammonia production on glutamine (KAmmGln) was 0.25 +/- 0.11 mM and 0.10 +/- 0.03 mM, at glucose concentrations of 0.28 mM and 5.6 mM respectively. At very low glucose concentrations, the glucose to lactate yield decreased markedly, showing a metabolic shift towards lower lactate production. This metabolic shift was also confirmed by the significant increase in the specific oxygen consumption rate also observed at low glucose concentrations. Although it was highly dependent on glucose concentration, the oxygen consumption also increased with the increase in glutamine concentration. At very low glutamine concentrations, the glutamine to ammonia yield increased, showing a more efficient glutamine metabolism.

Baby hamster kidney (BHK) cell aggregates grown in stirred vessels with different working volumes and impeller sizes were characterized. Using batch cultures, the range of agitation rates studied (25-100 rpm) led to aggregates with maximum sizes of 150 mum. Necrotic centers were not observed and cell specific productivity was independent of aggregate size. High cell viability was found for both single and adherent cells without an increase in cell death when agitation rate was increased. The increase in agitation rate affected aggregates by reducing their size and increasing their concentration and cell concentration in aggregates, while increasing the fraction of free cells in suspension. The experimental relationship between aggregate size and power dissipation rate per unit of mass was close to -1/4, suggesting a correlation with a critical turbulence microscale; this was independent of vessel scale and impeller geometry over the range investigated. Viscous stresses in the viscous dissipation subrange (below Kolmogoroff eddies) appear to be responsible for aggregate breakage. Under intense agitation BHK cells grown in the absence of microcarriers existed as aggregates without cell damage, whereas cells grown on the surface of microcarriers were largely reduced. This is a clear advantage for scaleup purposes if aggregates are used as a natural immobilization system in stirred vessels. (c) 1995 John Wiley & Sons, Inc.

RESUMOAmostras de queijo de minas artesanal foram coletadas em 18 queijarias localizadas em propriedades rurais da região da Serra da Canastra, Minas Gerais, com o objetivo de avaliar a influência da altitude sobre a população de bactérias acidolácticas. As queijarias estavam distribuídas nas altitudes de 600 a 900m, 900 a 1000m e mais de 1000m. Observaram-se populações mais elevadas de bactérias acidolácticas nas amostras de queijo da altitude de 600 a 900m. Lactobacillus rhamnosus, Lactobacillus casei e Lactobacillus plantarum foram os principais microrganismos isolados e identificados por PCR ARDRA 16S-23S rDNA, além de Enterococcus spp., Lactococcus spp. e outras espécies de Lactobacillus. Sugere-se que estas espécies estejam adaptadas ao ambiente de produção do queijo de minas artesanal produzido na região, o que resultaria em características sensoriais próprias do produto. Palavras-chave: queijo, altitude, bactérias acidolácticas, 16S-23S rDNA ABSTRACT Samples of minas artisanal cheese were collected in 18 small

Culture-dependent PCR-amplified rRNA gene restriction analysis and culture-independent (PCR-denaturing gradient gel electrophoresis) methodologies were used to examine vaginal lactobacilli from Brazilian women who were healthy or had been diagnosed with vulvovaginal candidiasis (VVC) or bacterial vaginosis. Only Lactobacillus crispatus was detected accordingly by both methods, and H 2 O 2 -producing lactobacilli were not associated with protection against VVC.