Regulation of carbon and electron flow in Propionispira arboris: physiological function of hydrogenase and its role in homopropionate formation

Abstract Hydrogenase activity was characterized in cell extracts of Propionispira arboris that consumed or produced H₂, coupled to methyl viologen reduction, and displayed highest levels (2.6 μmol/min/mg protein) in extracts prepared from fumarate-grown cells. Reversible hydrogenase activity in cell extracts correlated with the production of low levels of hydrogen during the growth phase and its subsequent consumption during the stationary phase of cells grown on glucose or lactate as the carbon and energy source. The addition of exogenous hydrogen to glucose, lactate or fumarate-grown cells dramatically increased propionate production at the expense of acetate formation. This accounted for the formation of propionate as nearly the sole end product of glucose fermentation under two atmospheres of hydrogen. The physiological function of hydrogenase in regulation of carbon and electron flow, and the significance of the results in applied and environmental microbiology are discussed.     

Decontamination of bacterial infection of monolayer cultures with a specific bacteriophage

Summary A few cell lines and primary monolayer cultures were accidentally infected by bacteria. These cultures were successfully decontaminated by means of the specific bacteriophage virus after quick identification of the responsible bacteria. This method presents a practical interest for preservation of valuable cultures. This work is supported by the Institut National de la Sante et de la Recherche Medicale (France) and the Fondation pour la Recherche Medicale (France).     

Assessment of environmental contamination associated with a mammalian cell transformation assay

Summary To estimate worker exposures to, and environmental contamination from, test chemicals and organic solvents used in an in vitro assay to assess the carcinogenic potential of chemicals, sodium fluorescein, a noncarcinogenic fluorescent material, was dissolved in tissue culture medium used to maintain early passage hamster embryo cells. Personal and environmental samples were taken over a 14-d period. The assay was performed according to standard procedures in a ventilated glove box or laminar flow safety cabinet. Considerably more than 99% of the chemical contamination found was recovered from the interiors of the glove box and hood and from disposable equipment. Contamination outside the containment units (less than 1 μg) resulted from intralaboratory transport of chemicals, treated cultures, and contaminated equipment. We conclude that the standard operating practices and procedures provided adequate safeguards for personnel and the environment. Research sponsored by the National Cancer Institute under Contract N01-CO-75380, with Litton Bionetics, Inc.     

Changes of intraocular pressure in patients with open-angle glaucoma in relation to the passage of atmospheric fronts and environmental contamination

Effects of atmospheric frontal passages on intraocular pressure (IOP) in patients with open-angle glaucoma are statistically investigated to show the meteorotropism of this disease. Changes in IOP caused by frontal passages are evident; the response is not identical in all the patients near the day of the passage of a warm front, while on the third day following the passage of the front a well pronounced drop in IOP occurs. Anomalous increases of IOP over several months' duration occurred in the years 1986–7. This finding is explained in relation to the hypothesis of environmental contamination in Central Europe by radioactive cesium nuclides due to the Chernobyl accident.     

Whole-lake experimentation as a tool to assess ecosystem health,response to stress and recovery: the Experimental Lakes Area experience

The rationale and methods for and value of whole-lake experimentation are described using the Experimental Lakes Area (ELA), northwestern Ontario, as the example. The ELA consists of 46 lakes (< 100 ha in surface area), their watersheds, and several streams protected for research purposes in near-pristine boreal forest on the Precambrian Shield near Kenora, Ontario. Over more than 20 y, whole-lake experimentation has provided unique information on the effects on lakes of nutrient additions, acidification, Cd addition, and biomanipulation. Experiments are planned to study the effects of PCB addition and flooding. Recovery, mitigation, and remediation have been explored in some experiments. As well, the fate of radioactive metals in a lake and the effects of acidification on a poor fen and an upland watershed are studied. Comparison between the experimental systems and unmanipulated reference systems has proven to be essential. These reference systems also have a role in defining (absolute) aquatic ecosystem health for small, pristine Precambrian Shield lakes. The ELA experimental data base is available, as well, for calibrating indices of relative aquatic ecosystem health, i.e., environmental degradation, using the dose-responses of lakes to eutrophication, acidification, Cd addition, and other stressors.     

Two relative efficiencies of polymorphic enzymes for characterizing cell lines,detecting contaminations,and monitoring transplants

Summary A new calculation of the relative efficiency of polymorphic enzyme markers, called the REB, was determined and compared with one of Fisher's determinations of the relative efficiency called REA here. The REA estimates the chance of failing, and 1-REA of succeeding, to show a phenotypic difference between two randomly selected persons or cultured cell lines (Case 1). In this study it was shown that the REA also estimates the chance of detecting a cell line mislabeling or similar mixup (Case 2) and a cell line cross-contamination leading to the complete replacement of an original line by contaminating line (Case 3). The new REB determines the probability of failing, and 1-REB of succeeding, to detect a contamination of an original line by another line leading to their coexistence, or at least a sufficiently long period of transitional coexistence before one overgrows the other. The REA and REB also apply to determining the efficiency of polymorphic markers in detecting donor and recipient cells in tissue transplants. This work was developed from the author's involvement in the human tumor cell-line characterization project at Sloan-Kettering Institute and he acknowledges this opportunity and the benefits of his association with Dr. J?rgen Fogh and colleagues in the Human Tumor Cell Laboratory.     

Separation of algal mixtures and bacterial mixtures with flow-microfluorometer using chlorophyll and ethidium bromide fluorescence

The applicability of flow-microfluorometer to separate microbial cells was demonstrated with algal and bacterial cells. Algal mixtures were sorted according to the natural chlorophyll fluorescence and the bacterial mixtures were sorted according to the fluorescence of ethidium bromide-stained nucleic acid.Abbreviation FMF Flow-microfluorometer     

Bovine pregnancy and nonreturn rates following artificial insemination using a covered sheath

Contamination transferred into the uterus from external genitalia during artificial insemination (AI) has been hypothesized to cause lowered bovine pregnancy rates (PR). Using aseptic techniques, there is still a possibility of uterine contamination during routine AI. Two experiments were conducted to evaluate the effect of two types of sheath covers (CS) placed over the conventional French Medium Syringe assembly (FMS) used for AI. Their use entailed passing the assembly to the external os of the cervix, pushing the FMS through the CS and manipulating the FMS to the cervical uterine junction in the normal manner. Fifty-six day non-return rate (NRR) in dairy and actual PR in beef cattle were evaluated. In Experiment 1, 30 professional technicians were employed to inseminate 7, 387 dairy cows, while in Experiment 2, six technicians with varying levels of experience inseminated 416 beef cows. Least-squares means for NRR in dairy cattle were 78% using a CS and 79% without. Means for PR in beef cattle were 57% using a CS and 62% without. In Experiment 2, the overall PR was lower in Trial l than in other trials (P<0.05). Since some technicians improved with time, the difference due to trial was attributed to technician variation in gaining experience with a CS. Results indicate that general use of a CS in routine AI of apparently healthy cows will not increase PR.     

Pitfalls in the sample preparation and analysis of N-acylethanolamines

N-acylethanolamines (NAEs) are a group of lipid mediators synthesized in response to a number of physiological and pathological stimuli. Because of the low tissue concentrations of NAEs, analyses often include liquid extraction followed by solid-phase extraction and subsequent quantitation by LC/MS or GC/MS. Reported levels of NAEs vary considerably, however, and often no explanation is given for these discrepancies. Brought on by difficulties encountered during method development, the effects of using four different brands of silica-containing solid phase extraction (SPE) columns and five different brands of chloroform for sample preparation were investigated. Considerable variation in the retention and recoveries of seven NAEs and 2-arachidonoylglycerol existed between the SPE columns. Furthermore, it was found that some chloroforms contained quantifiable amounts of N-palmitoylethanolamine and N-stearoylethanolamine. Finally, it was found that use of one of the chloroforms resulted in a loss of N-oleoylethanolamine from solution due to addition of chlorine to the ω-9 bond. The identity of this reaction product was confirmed by LC-MS/MS and NMR. It is recommended that these aspects of sample preparation and analysis should be thoroughly validated during method development and the relevant information on specific brands used be reported in future communications in order to better estimate the validity of reported quantitative data.     

Ground cover and slope exposure effects on micro- and mesobiota in forest soils

The interrelation of Alpine topography with the micro − and mesobiota is still poorly understood. We investigated the effects of ground cover type and slope exposure on the soil microbial biomass (double-stranded DNA, dsDNA) and abundances (real time PCR, qPCR); hydrolytic enzyme activities; and enchytraeid community structure in top soils (2.5-cm increments depth) in subalpine forests in the Italian Alps. Dominant ground covers were grass, moss, litter and woody debris at the north- and the south-facing slopes. The autochthonous soil microbiota (bacteria, fungi and archaea) was quantified by qPCR in the extracellular (eDNA) and intracellular fraction (iDNA) of the total soil DNA pool. A higher eDNA/iDNA ratio indicative of lower microbial activity was recorded in the deepest layer of the grass plots at the north-facing slope. This can be related to a lower degradation of eDNA and/or to an accumulation of eDNA with increasing depth as a result of leaching. The exposure effect was enzyme-specific and higher activities occurred under woody debris primarily at the south-facing slope. These plots also showed a higher nutrient content and a greater microbial biomass assessed as dsDNA yields. Total microannelid abundance was elevated on north-facing slopes on account of strong acidity indicator species. This was related to soil pH being one unit lower compared to the south-facing slope. The thickness of the organic layer (OL + OF + OH) was elevated at the north-facing slope due to a considerably thicker OH-horizon. The vast majority of microannelids at this slope occurred in the organic layer, while at south exposure they were almost evenly distributed between the organic layer and the mineral soil (A-horizon). Exposure was found to be more determinative for the composition of microannelid assemblages than the ground cover type.