Evaluation of a method for removing cesium and reducing the volume of leaf litter from broad-leaved trees contaminated by the Fukushima Daiichi nuclear accident during the Great East Japan Earthquake.

The town of Marumori in southern Miyagi Prefecture borders on Fukushima Prefecture, and following the accident at the Fukushima Daiichi nuclear power plant, there were concerns about cesium deposition in forested areas. One of the authors of this paper has continually surveyed leaf litter from the forested areas. As leaf litter may be a source of cesium contamination from the forest to downstream areas, we considered a simplified version of wet oxidation, a method previously presented by one of the authors of this study, as a technology to reduce leaf litter weight and cesium concentration, separating radioactive nuclides from non-radioactive ones, in leaf litter. We tested our method in three experiments. Experiment 1 used new leaf litter (232 Bq/kg) from the surface of a small stream at the forest edge nearby an area with air dose level higher than the national standard threshold of 0.23 µSv/h for the implementation of governmental decontamination works. Experiment 2 applied wet oxidation to older leaf litter (705 Bq/kg) harvested from a pasture nearby the stream mentioned above. We also used the same leaf litter in experiment 3 for a cesium release tests using pure water. In experiment 1 and 2 we treated leaf litter with a sodium hypochlorite solution, optimizing sodium hypochlorite concentration and reaction temperature. We measured a 50-60% decrease in the leaf litter weight and a 60% decrease in the cesium concentration. Moreover, we also measured the amount of cesium washout. The cesium budget of experiment 1 showed no cesium gasification (wet oxidation avoids airborne cesium as this element is prone to be volatile at 600 °C), and that high sodium hypochlorite concentration and high temperature had a strong positive effect on leaf litter volume reduction and cesium decontamination. Experiment 2 confirmed the reproducibility of these results in leaves with different cesium concentration and harvested in different conditions. We could also explain the mechanism behind leaf litter weight and cesium concentration reduction. Experiment 3 helped us to investigate the effects of the matter present on the surface of the water and the contribution of water soluble cesium. Concurrent experiments on changes in leaf litter chemical composition confirmed that our modified wet oxidation method had an effect on the removal of acid-insoluble lignin. Removal of lignin, a refractory component, might allow for a better utilization of the residue left after implementation of the proposed simplified wet oxidation. Thus, real wastes could be smaller than the residues. Together with the observed smaller cesium concentration in the residue, the proposed method in this study is expected to contribute to mitigate the risk due to the fallen leaves containing cesium.

Direct production of ethanol from neoagarobiose using recombinant yeast that secretes α-neoagarooligosaccharide hydrolase.

An α-neoagarooligosaccharide hydrolase, AgaNash, was purified from Cellvibrio sp. OA-2007, which utilizes agarose as a substrate. The agaNash gene, which encodes AgaNash, was obtained by comparing the N-terminal amino acid sequence of AgaNash with that deduced from the nucleotide sequence of the full-length OA-2007 genome. The agaNash gene combined with the Saccharomyces cerevisiae signal peptide α-mating factor was transformed into the YPH499 strain of S. cerevisiae to generate YPH499/pTEF-MF-agaNash, and the recombinant yeast was confirmed to produce AgaNash, though it was mainly retained within the recombinant cell. To enhance AgaNash secretion from the cell, the signal peptide was replaced with a combination of the signal peptide and a threonine- and serine-rich tract (T-S region) of the S. diastaticus STA1 gene. The new recombinant yeast, YPH499/pTEF-STA1SP-agaNash, was demonstrated to secrete AgaNash and hydrolyze neoagarobiose with an efficiency of as high as 84%, thereby producing galactose, which is a fermentable sugar for the yeast, and ethanol, at concentrations of up to 1.8 g/L, directly from neoagarobiose.

Draft Genome Sequence of the Nonmarine Agarolytic Bacterium Cellvibrio sp. OA-2007.

Cellvibrio sp. OA-2007 is a Gram-negative, aerobic, and agarolytic bacterium isolated from activated sludge. We present the draft genome sequence of strain OA-2007, composed of 97 contigs, totaling 4,595,379 bp in size, and containing 4,094 open reading frames, with a G+C content of 47.71%.

Strategies for the production of high concentrations of bioethanol from seaweeds: production of high concentrations of bioethanol from seaweeds.

Bioethanol has attracted attention as an alternative to petroleum-derived fuel. Seaweeds have been proposed as some of the most promising raw materials for bioethanol production because they have several advantages over lignocellulosic biomass. However, because seaweeds contain low contents of glucans, i.e., polysaccharides composed of glucose, the conversion of only the glucans from seaweed is not sufficient to produce high concentrations of ethanol. Therefore, it is also necessary to produce ethanol from other specific carbohydrate components of seaweeds, including sulfated polysaccharides, mannitol, alginate, agar and carrageenan. This review summarizes the current state of research on the production of ethanol from seaweed carbohydrates for which the conversion of carbohydrates to sugars is a key step and makes comparisons with the production of ethanol from lignocellulosic biomass. This review provides valuable information necessary for the production of high concentrations of ethanol from seaweeds.

Cloning of agarase gene from non-marine agarolytic bacterium Cellvibrio sp.

Agarase genes of non-marine agarolytic bacterium Cellvibrio sp. were cloned into Escherichia coli and one of the genes obtained using HindIII was sequenced. From nucleotide and putative amino acid sequences (713 aa, molecular mass; 78,771 Da) of the gene, designated as agarase AgaA, the gene was found to have closest homology to the Saccharophagus degradans (formerly, Microbulbifer degradans) 2-40 aga86 gene, belonging to glycoside hydrolase family 86 (GH86). The putative protein appears to be a non-secreted protein because of the absence of a signal sequence. The recombinant protein was purified with anion exchange and gel filtration columns after ammonium sulfate precipitation and the molecular mass (79 kDa) determined by SDS-PAGE and subsequent enzymography agreed with the estimated value, suggesting that the enzyme is monomeric. The optimal pH and temperature for enzymatic hydrolysis of agarose were 6.5 and 42.5 degrees C, and the enzyme was stable under 40 degrees C. LC-MS and NMR analyses revealed production of a neoagarobiose and a neoagarotetraose with a small amount of a neoagarohexaose during hydrolysis of agarose, indicating that the enzyme is a beta-agarase.

Microbiological quality of fermented milk produced by repeated-batch culture.

Using a starter culture containing Lactococcus lactis subsp. cremoris and Acetobacter orientalis, repeated-batch culture for fermented-milk production was carried out for 14 d. The pH decreased to approximately 4.3 and the milk solidified stably when the mixing ratio of fermented-milk to fresh milk was set at a level as high as 10%. The microbial population gradually changed as the culture progressed, and the cell densities of lactic and acetic acid bacteria finally leveled off to constant values. The quality of the fermented-milk became almost constant with respect to the changes in the cell densities of the lactic and acetic acid bacteria. Escherichia coli was then inoculated into the fermented-milk to simulate household contamination. E. coli was washed out with the progress of the repeated-batch culture, and thus showed no adverse effects on the fermented-milk production.