Geochemical and physiological evidence for mixed aerobic and anaerobic field biodegradation of coal tar waste by subsurface microbial communities

We used geochemical analyses of groundwater and laboratory-incubated microcosms to investigate the physiological responses of naturally occurring microorganisms to coal-tar-waste constituents in a contaminated aquifer. Waters were sampled from wells along a natural hydrologic gradient extending from uncontaminated (1 well) into contaminated (3 wells) zones. Groundwater analyses determined the concentrations of carbon and energy sources (pollutants or total organic carbon), final electron acceptors (oxygen, nitrate, sulfate), and metabolic byproducts (dissolved inorganic carbon [DIC], alkalinity, methane, ferrous iron, sulfide, Mn2+). In the contaminated zone of the study site, concentrations of methane, hydrogen, alkalinity, and DIC were enhanced, while dissolved oxygen and nitrate were depleted. Field-initiated biodegradation assays using headspace-free serum bottle microcosms filled with groundwater examined metabolism of the ambient organic contaminants (naphthalene, 2-methylnaphthalene, benzothiophene, and indene) by the native microbial communities. Unamended microcosms from the contaminated zone demonstrated the simultaneous degradation of several coal-tar-waste constituents at the in situ temperature (10°C). Lag phases prior to the onset of biodegradation indicated the prevalence of both aerobic and anaerobic conditions in situ. Electron acceptor-amended microcosms from the most contaminated well waters demonstrated only aerobic naphthalene degradation. Collectively, the geochemical and microbial evidence show that biodegradation of coal-tar-waste constituents occurs via both aerobic and anaerobic terminal electron accepting processes at this site.     

A plasmid selection system in Lactococcus lactis and its use for gene expression in L. lactis and human kidney fibroblasts

We report the development of a nonantibiotic and nonpathogenic host-plasmid selection system based on lactococcal genes and threonine complementation. We constructed an auxotrophic Lactococcus lactis MG1363Deltathr strain which carries deletions in two genes encoding threonine biosynthetic enzymes. To achieve plasmid-borne complementation, we then constructed the minimal cloning vector, pJAG5, based on the genes encoding homoserine dehydrogenase-homoserine kinase (the hom-thrB operon) as a selective marker. Using strain MG1363Deltathr, selection and maintenance of cells carrying pJAG5 were obtained in threonine-free defined media. Compared to the commonly used selection system based on erythromycin resistance, the designed complementation system offers a competitive and stable plasmid selection system for the production of heterologous proteins in L. lactis. The potential of pJAG5 to deliver genes for expression in eukaryotes was evaluated by insertion of a mammalian expression unit encoding a modified green fluorescent protein. The successful delivery and expression of genes in human kidney fibroblasts indicated the potential of the designed nonantibiotic host-plasmid system for use in genetic immunization.     

Propionic acid side chain hydrogen bonding in the malaria pigment beta-hematin

Malaria pigment, or beta-hematin, the insoluble heme detoxification product resulting from the intraerythrocitic digestion of hemoglobin by young malaria trophozoites has been structurally characterized by X-ray powder diffraction and shown to contain chains of propionic acid linked dimers. Although there is considerable spectroscopic evidence for a monodentate propionate-iron interaction in this crystalline material, the spectroscopic characterization of the propionic acid dimer is limited. Herein we demonstrate the presence of the propionic acid dimer unit by H/D isotope substitution in carboxylic acid dimer. In the Raman spectrum of the deuterium substituted compound there is a circa 12 cm(-1) shift, H: 1629 cm(-1) vs. D: 1617 cm(-1) in the symmetric ring breathing mode for the propionic acid dimer. On the other hand, the IR active asymmetric stretch has a very small shift, <3 cm(-1), upon deuteration. These, and other vibrational data, are consistent with the presence of a planar carboxylic acid dimer in the structure of beta-hematin.     

Cross-contamination with Salmonella on a broiler slaughterhouse line demonstrated by use of epidemiological markers

AIMS: To investigate contamination of surfaces on a poultry slaughter line from infected poultry and subsequent cross-contamination of non-infected poultry. METHODS AND RESULTS: A broiler slaughterhouse was investigated for the presence of Salmonella on 17 defined points over two 1-week periods. Flocks supplied to slaughter and neck skin samples from processed chicken were likewise investigated. Salmonella was detected in 10 out of 18 flocks at ante-mortem inspection, while seven flocks tested positive in the finished products. Equipment at all but one control point at the slaughter line tested positive at least once during the study. The chicken receiving area was the most contaminated. By comparison of typing results from serotyping, plasmid profile typing and phage typing, direct evidence for cross-contamination with Salm. serotype Typhimurium, Salm. Serotype 4.12:b:- and Salm. serotype Virchow on the slaughter line was obtained for four of the flocks. The cleaning procedure in place did not remove all Salmonella from the contaminated areas. CONCLUSIONS: Evidence for contamination of equipment on a slaughter line and subsequent cross-contamination to non-infected chicken was provided by typing methods. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has provided detailed information on cross-contamination on a slaughter line by the use of phage typing and plasmid profiling. The study stresses the importance of controlling Salmonella in the primary production, as contamination of the equipment on the slaughter line will act as a vehicle to contaminate finished products. Cleaning procedures on slaughter lines cannot be expected to control this problem with the current equipment.     

Chromosome location, genomic organization of the porcine COL10A1 gene and model structure of the NC1 domain

The porcine COL10A1 gene, encoding the alpha1(X) chain of type X collagen, has been sequenced. The gene structure is evolutionarily conserved, consisting of three exons and two introns spanning 7100 bp. Linkage mapping localized the gene to chromosome 1, which is in agreement with human-pig homology maps. Furthermore, protein structure comparison of the functionally important carboxyl domain between species revealed that amino acid changes were few and mainly situated in loop regions.     

NAGase activity in airborne biomass dust and relationship between NAGase concentrations and fungal spores

Inhalation of airborne fungal spores or fungalenzymes may cause adverse pulmonary healtheffects. The enzyme NAGase(N-acetyl--D-glucosaminidase) is achitinase presumed to be secreted by all fungi. In this study, NAGase activities andconcentrations of fungi are estimated inairborne biomass dust to acquire knowledgeabout the level of NAGase activity and therelationship between NAGase activity andconcentrations of airborne fungal spores.NAGase was sampled on both teflon andpolycarbonate filters, and polycarbonatefilters proved to be better for extraction ofNAGase than teflon filters. NAGase was foundin airborne dust at a biofuel plant and in dustgenerated from biomass. At a biofuel plant, themedian level of exposure to NAGase was 21 pmols^–1 m^–3. Significant correlationswere found between NAGase activities, totalnumber of fungal spores and CFU of fungi, withthe highest degree of correlation being betweenthe total number of fungal spores and theNAGase activity (r = 0.802, n = 76). Furthermore,when dust was stored for different periods, theculturability of fungal spores was stronglyreduced and the NAGase activity was not or onlyslightly reduced after up to 40 days ofstorage. Accordingly, NAGase activity may beused as a rapid method to get an estimate ofthe exposure level to airborne fungal spores.Whether pure NAGase or the NAGaseconcentrations observed here cause any healtheffects is not known, although it has beenshown that other fungal enzymes can causerespiratory disorders and a chitinase isdescribed as an allergen.     

High internal resistance to CO2 uptake by submerged macrophytes that use HCO3 −: measurements in air,nitrogen and helium

Previous studies have shown that in water the affinity of submerged macrophytes for CO₂ is higher for species restricted to CO₂ than for species with an additional ability to use bicarbonate. We measured slopes of CO₂ uptake versus CO₂ concentration in the gas phase in air, nitrogen and helium for pairs of species, having or lacking the ability to use bicarbonate, but with similar leaf morphology. For species restricted to CO₂, the slope in nitrogen and helium was 1.5 times and 3.2 times greater than in air. The increased slope in nitrogen results from a suppression of photorespiration. The further increase in helium reflects the increased rate of diffusion of CO₂ and shows that, even in gas, external diffusion through the boundary layer is a significant hindrance to CO₂ uptake. In contrast, in species able to use bicarbonate, the uptake slope was not affected by gas composition, suggesting that photorespiration is absent or photorespired CO₂ is efficiently trapped and that internal resistance is high relative to external resistance. Elodea canadensis specimens grown under high concentrations of CO₂ de-regulated their ability to use bicarbonate, and slopes of CO₂ uptake in helium were significantly greater than in air or nitrogen. Overall, these results are consistent with the notion that while a high affinity for CO₂ will maximise carbon uptake in species restricted to CO₂, for species able to use bicarbonate, a high internal resistance would reduce loss of CO₂ and help maintain high concentrations of CO₂ at the site of fixation. This revised version was published online in August 2006 with corrections to the Cover Date.