A hybrid anaerobic solid-liquid bioreactor for food waste digestion

A hybrid anaerobic solid-liquid (HASL) bioreactor is an enhanced two-phase anaerobic system, that consists of a solid waste reactor as the acidification reactor and a wastewater reactor, i.e. an upflow anaerobic sludge blanket (UASB) reactor as the methanogenic reactor. Food waste digestion in HASL bioreactors with pre-acidification and HASL operation stages was investigated in two separate runs. After 8 days of pre-acidification in Run A and 4 days in Run B, total volatile fatty acid (TVFA) and chemical oxygen demand (COD) concentrations in the leachates of both acidification reactors were similar. During HASL operation stage, TVFA and COD removal in the methanogenic phase were 77–100% and 75–95%, respectively. Some 99% of the total methane generated was from the methanogenic phase with a content of 68–70% methane. At the end of operation, about 59–60% of the added volatile solids (VS) were removed with a methane yield of 0.25 l g^–1 VS.     

Sex and hormonal modification of 6-keto-PGF1α release by rat aorta

The stable breakdown product of prostacyclin, 6-keto-PGF_1α, was estimated in plasma samples after incubation with rat aortic rings. The 6-keto-PGF_1α concentration obtained with the male aortae was two-fold higher than that of the female. Ovariectomy markedly increased 6-keto-PGF_1α six-fold, but castration had no effect. Estradiol and progesterone treatment of the ovariectomized female suppressed (by 50%) and enhanced (two-fold) 6-keto-PGF_1α. Testosterone was without effect in gonadectomized males and females. Castrate males did not respond to gonadal steroid treatment.     

The Acetyltransferase Activity of the Bacterial Toxin YopJ of Yersinia Is Activated by Eukaryotic Host Cell Inositol Hexakisphosphate

Plague, one of the most devastating diseases in human history, is caused by the bacterium Yersinia pestis. The bacteria use a syringe-like macromolecular assembly to secrete various toxins directly into the host cells they infect. One such Yersinia outer protein, YopJ, performs the task of dampening innate immune responses in the host by simultaneously inhibiting the MAPK and NFκB signaling pathways. YopJ catalyzes the transfer of acetyl groups to serine, threonine, and lysine residues on target proteins. Acetylation of serine and threonine residues prevents them from being phosphorylated thereby preventing the activation of signaling molecules on which they are located. In this study, we describe the requirement of a host-cell factor for full activation of the acetyltransferase activity of YopJ and identify this activating factor to be inositol hexakisphosphate (IP₆). We extend the applicability of our results to show that IP₆ also stimulates the acetyltransferase activity of AvrA, the YopJ homologue from Salmonella typhimurium. Furthermore, an IP₆-induced conformational change in AvrA suggests that IP₆ acts as an allosteric activator of enzyme activity. Our results suggest that YopJ-family enzymes are quiescent in the bacterium where they are synthesized, because bacteria lack IP₆; once injected into mammalian cells by the pathogen these toxins bind host cell IP₆, are activated, and deregulate the MAPK and NFκB signaling pathways thereby subverting innate immunity.     

Virus harvesting in perfusion culture: Choosing the right type of hollow fiber membrane

The use of bioreactors coupled to membrane-based perfusion systems enables very high cell and product concentrations in vaccine and viral vector manufacturing. Many virus particles, however, are not stable and either lose their infectivity or physically degrade resulting in significant product losses if not harvested continuously. Even hollow fiber membranes with a nominal pore size of 0.2 µm can retain much smaller virions within a bioreactor. Here, we report on a systematic study to characterize structural and physicochemical membrane properties with respect to filter fouling and harvesting of yellow fever virus (YFV; ~50 nm). In tangential flow filtration perfusion experiments, we observed that YFV retention was only marginally determined by nominal but by effective pore sizes depending on filter fouling. Evaluation of scanning electron microscope images indicated that filter fouling can be reduced significantly by choosing membranes with (i) a flat inner surface (low boundary layer thickness), (ii) a smooth material structure (reduced deposition), (iii) a high porosity (high transmembrane flux), (iv) a distinct pore size distribution (well-defined pore selectivity), and (v) an increased fiber wall thickness (larger effective surface area). Lowest filter fouling was observed with polysulfone (PS) membranes. While the use of a small-pore PS membrane (0.08 µm) allowed to fully retain YFV within the bioreactor, continuous product harvesting was achieved with the large-pore PS membrane (0.34 µm). Due to the low protein rejection of the latter, this membrane type could also be of interest for other applications, that is, recombinant protein production in perfusion cultures.     

A straightforward radiometric technique for measuring IMP dehydrogenase

[2-3H]Inosinic acid ([2-3H]IMP) has been biosynthesized in good yield from [2-3H]hypoxanthine and PRPP via the action of a partially purified preparation of hypoxanthine/guanine phosphoribosyl transferase from mouse brain. The product was purified in one step by ascending paper chromatography, and used to assess the activity of IMP dehydrogenase. To conduct the assay, tritiated substrate is admixed with enzyme in a final volume of 10 microliters; NAD is present to serve as cofactor for the reaction, and allopurinol to inhibit the oxidation of any hypoxanthine generated as a consequence of side reactions. After an appropriate period of incubation, the 3H2O arising from the oxidation of tritiated IMP via [3H]NAD is isolated by quantitative microdistillation. Performed as described, the assay is facile, sensitive, and accurate, with the capability of detecting the dehydrogenation of as little as 1 pmol of [3H]IMP. Using it, measurements have been made of IMP dehydrogenase in a comprehensive array of mouse organs. Of these, pancreas contained the enzyme at the highest specific activity.     

Observations of a set of isomeric anti-lactose antibodies directed against a group D streptococcal polysaccharide

Sets of isomeric anti-lactose antibodies with specificity for the lactose units of a cell wall polysaccharide fromStreptococcus faecalis strain N were induced in rabbits immunized with a vaccine of nonviable cells of the organism. Such sets of anti-lactose antibodies were isolated from the serum of immunized animals by affinity chromatography on lactosyl-Sepharose. Gel electrofocusing experiments showed that the preparations consisted of multiprotein components. One preparation of antibodies of 13 isomers was separated into homogeneous components by liquid isoelectrofocusing. The individual isomeric antibodies exhibit specificity for the lactose units of the antigenic polysaccharide, possess isoelectric points in the range of 5.9–8.0, and belong to the IgG class of immunoglobulins, and each member yields one light chain and one heavy chain on dissociation in sodium dodecyl sulfate (SDS) and mercaptoethanol. These results have been interpreted as evidence for the assembly of the chains of isomeric antibodies by a single-chain pairing mechanism.     

Partial ischemia and proteinuria during isolated kidney perfusion is accompanied by the release of vascular [35S]heparan sulfate

The isolated kidney perfused with modified Krebs-Henseleit buffer with amino acids yields heavy proteinuria associated with reduced ATP levels characteristic of partial ischemia. These conditions are associated with a similar perfusion time dependent release of degraded vascular [35S]heparan sulfate proteoglycan into the perfusate solution which included a 60% loss of [35S]macromolecular material from the glomerulus after 2h of perfusion. Small amounts of [35S]macromolecular material were found in the urine and lymph. These results demonstrate that partial ischemia promotes a specific response in the overall renal vasculature, probably involving oxygen reactive metabolites, that results in the preferential release of heparan sulphate from the basement membrane and endothelial cells on the luminal side of the capillary wall.