Effect of heat-alkaline treatment as a pretreatment method on volatile fatty acid production and protein degradation in excess sludge, pure proteins and pure cultures.

This study investigated the effect of heat-alkaline treatment (HAT) at pH 11 and 60 °C on volatile fatty acid (VFA) production and protein degradation in excess sludge, soluble and insoluble proteins, and pure cultures. In addition, quantification of bacteria present in the sludge was also examined. Experimental results showed that following acid fermentation under pH 7 and 37 °C, HAT enhanced VFA production in excess sludge, albumin, and Gram-negative bacteria, but not in casein or Gram-positive bacteria. Protein solubility was therefore found not to be the main criteria for VFA production. In the protein analysis, it was shown that the outer membrane protein (OmpC) of Escherichia coli K12 was resistant to chemical and enzymatic hydrolysis. Gram staining revealed that Gram-negative bacteria were predominant in the activated sludge used in this study. In addition, the bacteria present in the activated sludge comprised only 10% of mixed liquor suspended solids (MLSS) by quantitative PCR.

Sustainable engineered processes to mitigate the global arsenic crisis in drinking water: challenges and progress.

Millions of people around the world are currently living under the threat of developing serious health problems owing to ingestion of dangerous concentrations of arsenic through their drinking water. In many places, treatment of arsenic-contaminated water is an urgent necessity owing to a lack of safe alternative sources. Sustainable production of arsenic-safe water from an arsenic-contaminated raw water source is currently a challenge. Despite the successful development in the laboratory of technologies for arsenic remediation, few have been successful in the field. A sustainable arsenic-remediation technology should be robust, composed of local resources, and user-friendly as well as must attach special consideration to the social, economic, cultural, traditional, and environmental aspects of the target community. One such technology is in operation on the Indian subcontinent. Wide-scale replication of this technology with adequate improvisation can solve the arsenic crisis prevalent in the developing world.

Cell envelope fluidity modification for an effective glutamate excretion in Corynebacterium glutamicum 2262.

1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) was used to assess the cell envelope fluidity of Corynebacterium glutamicum 2262 during a temperature-triggered glutamate producing process. Because the fluorescence lifetime of TMA-DPH was shown to be constant all over the process, fluorescence anisotropy can be considered as a good index of cell envelope fluidity. When the temperature of the fed-batch culture was increased from 33 to 39 degrees C to induce glutamate excretion, the fluorescence anisotropy values decreased from 0.212 +/- 0.002 to 0.186 +/- 0.002 (corresponding to an increase in the cell fluidity), while the specific glutamate production rate reached its maximal value. The increase in fluidity of the C. glutamicum cell envelope was not due to a physical effect related to the temperature elevation, but rather to an alteration of the composition of the cell envelope. Using a mutant devoid of corynomycolates, significant differences in fluorescence anisotropy values were obtained compared to the wild-type strain, suggesting that TMA-DPH is mainly anchored into the corynomycomembrane. Differences in fluorescence anisotropy were also observed when the bacteria were cultivated at 33, 36, 38, and 39 degrees C in batch cultures, and a linear relationship was obtained between the maximum specific glutamate production rate and the measured fluidity. When using the glutamate non-producing variant of C. glutamicum 2262, the fluorescence anisotropy remained constant at 0.207 +/- 0.003 whatever the applied temperature shift. This suggests that the fluidity of the Corynebacteria mycomembrane plays an important role in glutamate excretion during the temperature-triggered process.

Characterization of a Corynebacterium glutamicum lactate utilization operon induced during temperature-triggered glutamate production.

Gene expression changes of glutamate-producing Corynebacterium glutamicum were identified in transcriptome comparisons by DNA microarray analysis. During glutamate production induced by a temperature shift, C. glutamicum strain 2262 showed significantly higher mRNA levels of the NCgl2816 and NCgl2817 genes than its non-glutamate-producing derivative 2262NP. Reverse transcription-PCR analysis showed that the two genes together constitute an operon. NCgl2816 putatively codes for a lactate permease, while NCgl2817 was demonstrated to encode quinone-dependent l-lactate dehydrogenase, which was named LldD. C. glutamicum LldD displayed Michaelis-Menten kinetics for the substrate l-lactate with a K(m) of about 0.51 mM. The specific activity of LldD was about 10-fold higher during growth on l-lactate or on an l-lactate-glucose mixture than during growth on glucose, d-lactate, or pyruvate, while the specific activity of quinone-dependent d-lactate dehydrogenase differed little with the carbon source. RNA levels of NCgl2816 and lldD were about 18-fold higher during growth on l-lactate than on pyruvate. Disruption of the NCgl2816-lldD operon resulted in loss of the ability to utilize l-lactate as the sole carbon source. Expression of lldD restored l-lactate utilization, indicating that the function of the permease gene NCgl2816 is dispensable, while LldD is essential, for growth of C. glutamicum on l-lactate.

Dynamics of glutamate synthesis and excretion fluxes in batch and continuous cultures of temperature-triggered Corynebacterium glutamicum.

Corynebacterium glutamicum 2262 strain, when triggered for glutamate excretion, experiences a rapid decrease in growth rate and increase in glutamate efflux. In order to gain a better quantitative understanding of the factors controlling the metabolic transition, the fermentation dynamics was investigated for a temperature-sensitive strain cultivated in batch and glucose-limited continuous cultures. For non-excreting cells at 33 degrees C, increasing the growth rate resulted in strong increases in the central metabolic fluxes, but the intracellular glutamate level, the oxoglutarate dehydrogenase complex (ODHC) activity and the flux distribution at the oxoglutarate node remained essentially constant. When subjected to a temperature rise to 39 degrees C, at both high- and low-metabolic activities, the bacteria showed a rapid attenuation in ODHC activity and an increase from 28% to more than 90% of the isocitrate dehydrogenase flux split towards glutamate synthesis. Simultaneously to the reduction in growth rate, the cells activated a high capacity export system capable of expelling the surplus of synthesized glutamate.

Instability of glutamate production by Corynebacterium glutamicum 2262 in continuous culture using the temperature-triggered process.

Kinetics and physiology of Corynebacterium glutamicum 2262 cultured for extended periods in continuous mode were investigated at 33, 39 and 41 degrees C. At 33 degrees C no glutamate production occurred whatever the dilution rates tested (ranging between 0.05 and 0.5 h(-1)). When the continuous culture was performed at 39 degrees C and D=0.05 h(-1), the glutamate was actively produced, while the activities of 2-oxoglutarate dehydrogenase complex (ODHC) and pyruvate dehydrogenase (PDH) were, respectively completely inhibited and 35% decreased. Simultaneously, the intracellular glutamate was 62% reduced compared to the level found at 33 degrees C and the co-metabolites lactate and trehalose were excreted. The decrease in PDH activity during the glutamate production was suggested to be responsible for the accumulation of by-products and for limiting the carbon flux required for glutamate synthesis. When the culture was prolonged for more than 100 h, a cell selection occurred, in favor of growth and to the detriment of glutamate production. In fact, these selected cells presented high levels of ODHC and PDH activities even at 39 degrees C, resulting in a complete inhibition of the glutamate production after 150 h of culture. A further temperature increase till 41 degrees C restored the glutamate production and abolished the ODHC activity of these selected cells.