Iodate and nitrate transformation by Agrobacterium/Rhizobium related strain DVZ35 isolated from contaminated Hanford groundwater.

Nitrate and radioiodine (129I) contamination is widespread in groundwater underneath the Central Plateau of the Hanford Site. 129I, a byproduct of nuclear fission, is of concern due to a 15.7 million year half-life, and toxicity. The Hanford 200 West Area contains plumes covering 4.3 km2 with average 129I concentrations of 3.5 pCi/L. Iodate accounts for 70.6% of the iodine present and organo-iodine and iodide make up 25.8% and 3.6%, respectively. Nitrate plumes encompassing the 129I plumes have a surface area of 16 km2 averaging 130 mg/L. A nitrate and iodate reducing bacterium closely related to Agrobacterium, strain DVZ35, was isolated from sediment incubated in a 129I plume. Iodate removal efficiency was 36.3% in transition cultures, and 47.8% in anaerobic cultures. Nitrate (10 mM) was also reduced in the microcosm. When nitrate was spiked into the microcosms, iodate removal efficiency was 84.0% and 69.2% in transition and anaerobic cultures, respectively. Iodate reduction was lacking when nitrate was absent from the growth medium. These data indicate there is simultaneous reduction of nitrate and iodate by DVZ35, and iodate is reduced to iodide. Results provide the scientific basis for combined nitrogen and iodine cycling throughout the Hanford Site.

Microbial bioproducts from cheese whey through fermentation with wastewater sludge Clostridium isolates.

We demonstrated the production of hydrogen, ethanol, and a variety of acids by several Clostridium species using cheese whey as substrate. These species were isolated from the anaerobic sediments of a municipal wastewater stabilization pond. Eight isolates were obtained and all were classified taxonomically as Clostridium spp. based on 16S rRNA sequencing. Sludge isolates showed maximum bioproduct production yields and productivities after approximately 24 h of batch cultivation with 6% (m/v) cheese whey. Fermentation byproducts measured included hydrogen, ethanol, acetic acid, butyric acid, and lactic acid. The maximum yields of bioproducts were 0.59 mol H(2)/mol lactose, 0.071 g ethanol/g, 0.204 g acetic acid/g, 0.218 g butyric acid/g, and 0.144 g lactic acid/g. The production of these high value biofuels and biofuel intermediates from cheese whey could have significant implications for conversion of waste to high value bioproducts to enhance domestic energy economies.

Effect of coagulant/flocculants on bioproducts from microalgae.

The potential of microalgae as a source of sustainable energy, nutritional supplements and specialized chemicals necessitates a thorough evaluation of the methods of harvesting microalgae with regards to the bioproduct(s) desired. This research assessed the effect of coagulation, flocculation, and centrifugation on the wet lipid extraction procedure, which fractionated microalgae into hydrolyzed biomass for fermentation into acetone, butanol, and ethanol, an aqueous phase as growth media for genetically engineered Escherichia coli, and a lipid fraction for the production of biodiesel. Biomass harvested by cationic starches, alum, and centrifugation produced 30, 19, and 22.5mg/g of dry wt. algae of total combined acetone, butanol, and ethanol, respectively. Higher biodiesel production was also observed for the cationic starches (9.6 mg/g of dry wt. algae) than alum (0.6 mg/g of dry wt. algae) harvested biomass. The results suggested significant effect of the harvesting methods on the yields of bioproducts.

Monitoring microbial diversity of bioreactors using metagenomic approaches.

With the rapid development of molecular techniques, particularly 'omics' technologies, the field of microbial ecology is growing rapidly. The applications of next generation sequencing have allowed researchers to produce massive amounts of genetic data on individual microbes, providing information about microbial communities and their interactions through in situ and in vitro measurements. The ability to identify novel microbes, functions, and enzymes, along with developing an understanding of microbial interactions and functions, is necessary for efficient production of useful and high value products in bioreactors. The ability to optimize bioreactors fully and understand microbial interactions and functions within these systems will establish highly efficient industrial processes for the production of bioproducts. This chapter will provide an overview of bioreactors and metagenomic technologies to help the reader understand microbial communities, interactions, and functions in bioreactors.

Acetone, butanol, and ethanol production from wastewater algae.

Acetone, butanol, and ethanol (ABE) fermentation by Clostridium saccharoperbutylacetonicum N1-4 using wastewater algae biomass as a carbon source was demonstrated. Algae from the Logan City Wastewater Lagoon system grow naturally at high rates providing an abundant source of renewable algal biomass. Batch fermentations were performed with 10% algae as feedstock. Fermentation of acid/base pretreated algae produced 2.74 g/L of total ABE, as compared with 7.27 g/L from pretreated algae supplemented with 1% glucose. Additionally, 9.74 g/L of total ABE was produced when xylanase and cellulase enzymes were supplemented to the pretreated algae media. The 1% glucose supplement increased total ABE production approximately 160%, while supplementing with enzymes resulted in a 250% increase in total ABE production when compared to production from pretreated algae with no supplementation of extraneous sugar and enzymes. Additionally, supplementation of enzymes produced the highest total ABE production yield of 0.311 g/g and volumetric productivity of 0.102 g/Lh. The use of non-pretreated algae produced 0.73 g/L of total ABE. The ability to engineer novel methods to produce these high value products from an abundant and renewable feedstock such as algae could have significant implications in stimulating domestic energy economies.

Characterization of a Methanogenic Community within an Algal Fed Anaerobic Digester.

The microbial diversity and metabolic potential of a methanogenic consortium residing in a 3785-liter anaerobic digester, fed with wastewater algae, was analyzed using 454 pyrosequencing technology. DNA was extracted from anaerobic sludge material and used in metagenomic analysis through PCR amplification of the methyl-coenzyme M reductase α subunit (mcrA) gene using primer sets ML, MCR, and ME. The majority of annotated mcrA sequences were assigned taxonomically to the genera Methanosaeta in the order Methanosarcinales. Methanogens from the genus Methanosaeta are obligate acetotrophs, suggesting this genus plays a dominant role in methane production from the analyzed fermentation sample. Numerous analyzed sequences within the algae fed anaerobic digester were unclassified and could not be assigned taxonomically. Relative amplicon frequencies were determined for each primer set to determine the utility of each in pyrosequencing. Primer sets ML and MCR performed better quantitatively (representing the large majority of analyzed sequences) than primer set ME. However, each of these primer sets was shown to provide a quantitatively unique community structure, and thus they are of equal importance in mcrA metagenomic analysis.

Thermostable and Alkalistable Xylanases Produced by the Thermophilic Bacterium Anoxybacillus flavithermus TWXYL3.

With the rising cost and finite supply of fossil energy, there is an increasing economic incentive for the development of clean, efficient, and renewable domestic energy. The activities of microorganisms offer the potential conversion of lignocellulosic materials into fermentable sugars, usable for downstream fermentation processes. Strain TWXYL3, a thermophilic facultative anaerobe, was discovered in the Alvord Basin hydrothermal system in Oregon, USA. Phylogenetic analysis of strain TWXYL3 showed it to be 99% similar to the 16S rRNA gene of Anoxybacillus flavithermus WL (FJ950739). A. flavithermus TWXYL3 was shown to secrete a large multisubunit thermostable xylanase complex into the growth medium. Xylanase induction was achieved by resuspending the isolate in a selective xylan-containing medium. Extracellular xylanase activity showed a temperature optimum of 65°C and retained thermostability up to 85°C. Extracellular xylanase activity showed a bimodal pH optimum, with maxima at pH 6 and pH 8. Electrophoretic analysis of the extracellular xylanase shows 5 distinct proteins with xylanase activity. Strain TWXYL3 is the first xylanolytic isolate obtained from the Alvord Basin hydrothermal system and represents a new model system for development of processes where lignocellulosics are converted to biofuel precursors.