Lamb survival,glutathione redox state and immune function of neonates and lambs from periparturient Merino ewes supplemented with rumen-protected methionine

Wool growth in Merino sheep demands a high level of sulphur amino acids, competing with body growth and the immune system, which may play a role in increasing the risk of lamb mortality. The hypothesis that dietary supplementation of methionine (Met) to Merino ewes during the late stages of pregnancy will improve foetal growth and alter immune competency of ewes and lambs was tested in a total of 120 grazing, pregnant Merino ewes. Sixty ewes were group-supplemented with 6.3 g/d rumen-protected Met (Met-Plus) per sheep from day 111 of pregnancy until day 7 after lambing, and the other 60 animals were used as a non-supplemented Control. Lambs from Met-supplemented ewes tended to be 10% heavier than Control lambs (p = 0.10), which did not affected the survival rate at weaning significantly. The supplemented ewes had slightly higher concentrations of total glutathione (GSH) in plasma at lambing (p < 0.06), but there were no differences between 1 and 3 months post-lambing in GSH, glutathione disulphide (GSSG) and the GSSG:GSH ratio. The GSSG:GSH ratio in the blood of ewes was elevated at lambing (p < 0.05), hinting that ewes were undergoing increased oxidative stress. The Met supplementation elevated the total IgG concentration (p < 0.05) in lambs aged 4 and 6 weeks, but did not change the IgG concentrations in colostrum and in plasma of 1-week-old lambs, and white blood cell counts and leukocyte types. The trend towards higher lamb birth weights in the Met-supplemented group requires further investigation as this may influence survival at birth and weaning.     

C2 and C3 hydrocarbon gases associated with highly reducing conditions in groundwater

This study investigates the presence and concentration of light hydrocarbon gases in soil vapor located immediately above the capillary fringe of a petroleum-contaminated aquifer. A correlation was observed for the linear regression plot of redox potential versus detectable C₂+C₃ alkane concentrations for a limited number of sampling points. C₂+C₃ alkanes were not detected at points were redox potentials in groundwater exceeded -260 millivolts. The predominance of methanogenic processes in this redox range, as well as the observed C₂+C₃ concentration ratios, suggest that ethane and propane gases in soil vapor may be biogenically produced as well as a result of volatilization from NAPL.     

Impact of iron particles in groundwater on the UV inactivation of bacteriophages MS2 and T4

AIMS: To investigate the impact of iron particles in groundwater on the inactivation of two model viruses, bacteriophages MS2 and T4, by 254-nm ultraviolet (UV) light. METHODS AND RESULTS: One-litre samples of groundwater with high iron content (from the Indianapolis Water Company, mean dissolved iron concentration 1.3 mg l(-1)) were stirred vigorously while exposed to air, which oxidized and precipitated the dissolved iron. In parallel samples, ethylenediaminetetra-acetic acid (EDTA) was added to chelate the iron and prevent formation of iron precipitate. The average turbidity in the samples without EDTA (called the 'raw' samples) after 210 min of stirring was 2.7 +/- 0.1 NTU while the average turbidity of the samples containing EDTA (called the 'preserved' samples) was 1.0 +/- 0.1 NTU. 'Raw' and 'preserved' samples containing bacteriophage MS2 were exposed to 254-nm UV light at doses of 20, 40, or 60 mJ (cm(2))(-1), while samples containing bacteriophage T4 were exposed to 2 or 5 mJ (cm(2))(-1), using a low pressure UV collimated beam. The UV inactivation of both phages in the 'raw' groundwater was lower than in the EDTA-'preserved' groundwater to a statistically significant degree (alpha = 0.05), due to the association of phage with the UV-absorbing iron precipitate particles. A phage elution technique confirmed that a large fraction of the phage that survived the UV exposures were particle-associated. CONCLUSIONS: Phages that are associated with iron oxide particles in groundwater are shielded from UV light to a measurable and statistically significant degree at a turbidity level of 2.7 NTU when the phage particle association is induced under experimental conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: While the particle association of the phage in this study was induced experimentally, the findings provide further evidence that certain particles in natural waters and wastewaters (e.g. iron oxide particles) may have the potential to shield viruses from UV light.     

Assimilation of CO2 and introduced organic compounds by bacterial communities in groundwater from southeastern Sweden deep crystalline bedrock

The nutritional responses of unattached and attached bacterial communities were studied in groundwater from 3 sampling depths, i.e., 830–841 m, 910–921 m, and 999–1,078 m, of the subvertical borehole KLX01 at the Laxemar study area in SE Sweden. The salinity profile of the groundwater in this borehole is homogeneous. There were negative redox potentials (Eh) in the waters (–220 to –270 mV) and they contained sulfide, hydrogen, and methane. Biofilm reactors with hydrophilic glass surfaces were connected to the flowing groundwaters from each of the 3 depths with flow rates of approximately 3 x 10^–3 m sec^–1 over 19 days. There were 0.15 to 0.68 × 10⁵ unattached bacteria ml^–1 groundwater and 0.94 to 1.2 × 10⁵ attached bacteria cm^–2 on the surfaces. The assimilations of ¹⁴CO₂, ¹⁴C-formate, 1,2,3-³H-acetate, U-¹⁴C-lactate, U-¹⁴C-glucose, and L-4,5-³H-leucine by the communities were demonstrated with microautoradiographic and liquid scintillation counting techniques. There were significant assimilations of CO₂ by all communities, except for the unattached bacteria at the 910–921 m depth, indicating in situ production of organic carbon from carbonate. Assimilation of formate was detected in two communities, indicating the presence of bacteria able to substitute CO₂ with formate. Acetate, lactate, and glucose assimilations demonstrated the presence of heterotrophic bacteria. The assimilation of lactate by the attached bacteria dominated over acetate and glucose at all depths. Leucine was assimilated by 20 to 98% of the communities, which showed that major portions of the communities studied were viable. The results indicate that the attached communities at the 830–841 m and 910–921 m depths were in more metabolically active states than the unattached bacteria. Incubation in air compared with N₂ indicated that portions of the studied communities were obligate anaerobes, as their ability to assimilate the added compounds was sensitive to oxygen. The results show that the use of several different compounds reduces the risk for false conclusions about the viability and the metabolic activity of the deep groundwater communities.Offprint requests to: K. Pedersen.     

Impacts on microbial communities and cultivable isolates from groundwater contaminated with high levels of nitric acid-uranium waste

Microbial communities were characterized at contaminated sites that had elevated levels of nitrate, nickel, aluminum, and uranium (up to 690 mM, 310 microM, 42 mM, and 30 microM, respectively). The bacterial community structure based upon clonal libraries of the SSU rRNA genes (screened clones = 876) was diverse at the background site, but the three acidic samples had decreased diversity and the majority of clones were closely related to Azoarcus and Pseudomonas species. Arthrobacter and Novosphingobium sequences were recovered from the background samples but not the acidic sites, and similar pseudomonad populations were present at the background and acidic sites albeit at different relative abundances. Heterologous sequence coverage analyses indicated the microbial communities at the contaminated sites were very similar (p = 0.001) but different from the background site. Bacterial isolates (n = 67) classified as beta-or gamma-Proteobacteria, high G+C Gram-positive or low G+C Gram-positive were obtained from the background and one contaminated sample, and some of the isolates had less than 95% sequence identity with previously observed microorganisms. Despite variations in nitrate and heavy metal levels and different proximities to the source ponds, the three acidic samples had similar microbial populations. However, the least contaminated site (lowest nitrate and aluminum) had increased diversity compared to the other acidic samples. The results suggested that the combined contamination has decreased the microbial diversity, and Azoarcus populations were observed at a drastically increased frequency compared to the background site that had a more even distribution of multiple taxa.     

Biogeochemistry and microbial ecology of methane oxidation in anoxic environments: a review

Evidence supporting a key role for anaerobic methane oxidation in the global methane cycle is reviewed. Emphasis is on recent microbiological advances. The driving force for research on this process continues to be the fact that microbial communities intercept and consume methane from anoxic environments, methane that would otherwise enter the atmosphere. Anaerobic methane oxidation is biogeochemically important because methane is a potent greenhouse gas in the atmosphere and is abundant in anoxic environments. Geochemical evidence for this process has been observed in numerous marine sediments along the continental margins, in methane seeps and vents, around methane hydrate deposits, and in anoxic waters. The anaerobic oxidation of methane is performed by at least two phylogenetically distinct groups of archaea, the ANME-1 and ANME-2. These archaea are frequently observed as consortia with sulfate-reducing bacteria, and the metabolism of these consortia presumably involves a syntrophic association based on interspecies electron transfer. The archaeal member of a consortium apparently oxidizes methane and shuttles reduced compounds to the sulfate-reducing bacteria. Despite recent advances in understanding anaerobic methane oxidation, uncertainties still remain regarding the nature and necessity of the syntrophic association, the biochemical pathway of methane oxidation, and the interaction of the process with the local chemical and physical environment. This review will consider the microbial ecology and biogeochemistry of anaerobic methane oxidation with a special emphasis on the interactions between the responsible organisms and their environment. This revised version was published online in August 2006 with corrections to the Cover Date.     

Taxonomically and functionally diverse microbial communities in deep crystalline rocks of the Fennoscandian shield

Microbial life in the nutrient-limited and low-permeability continental crystalline crust is abundant but remains relatively unexplored. Using high-throughput sequencing to assess the 16S rRNA gene diversity, we found diverse bacterial and archaeal communities along a 2516-m-deep drill hole in continental crystalline crust in Outokumpu, Finland. These communities varied at different sampling depths in response to prevailing lithology and hydrogeochemistry. Further analysis by shotgun metagenomic sequencing revealed variable carbon and nutrient utilization strategies as well as specific functional and physiological adaptations uniquely associated with specific environmental conditions. Altogether, our results show that predominant geological and hydrogeochemical conditions, including the existence and connectivity of fracture systems and the low amounts of available energy, have a key role in controlling microbial ecology and evolution in the nutrient and energy-poor deep crustal biosphere.     

Metabolic and phylogenetic analysis of microbial communities during phytoremediation of soil contaminated with weathered hydrocarbons and heavy metals

In the current study, the microbial ecology of weathered hydrocarbon and heavy metal contaminated soil undergoing phytoremediation was studied. The relationship of functional diversity, measured as carbon source utilisation in Biolog plates and extracellular enzymatic activities, and genetic diversity of bacteria was evaluated. Denaturing gradient gel electrophoresis was used for community analyses at the species level. Bulk soil and rhizosphere soil from pine and poplar plantations were analysed separately to determine if the plant rhizosphere impacted hydrocarbon degradation. Prevailing microbial communities in the field site were both genetically and metabolically diverse. Furthermore, both tree rhizosphere and fertilisation affected the compositions of these communities and increased activities of extracellular aminopeptidases. In addition, the abundance of alkane hydroxylase and naphthalene dioxygenase genes in the communities was low, but the prevalence of these genes was increased by the addition of bioavailable hydrocarbons. Tree rhizosphere communities had greater hydrocarbon degradation potential than those of bulk soil. Hydrocarbon utilising communities were dominated generally by the species Ralstonia eutropha and bacteria belonging to the genus Burkholderia. Despite the presence of viable hydrocarbon-degrading microbiota, decomposition of hydrocarbons from weathered hydrocarbon contaminated soil over four years, regardless of the presence of vegetation, was low in unfertilised soil. Compost addition enhanced the removal of hydrocarbons.     

Optimisation of the dehydrogenase assay for measurement of indigenous microbial activity in beach sediments contaminated with petroleum

A dehydrogenase enzyme assay has been developed to measure microbial activity in petroleum-contaminated beach sediments. The assay, using 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazoliumchloride (INT) as the substrate provides a reliable measurement of overall indigenous microbiological activity in beach sediments containing 7% (w/w) oil. The method has been effectively used in field studies (Mathew et al. 1999) to monitor microbial activity in conjunction with the biodegradation of petroleum hydrocarbons.     

Campylobacterota dominate the microbial communities in a tropical karst subterranean estuary,with implications for cycling and export of nitrogen to coastal waters

Subterranean estuaries (STEs), the zones in which seawater and subsurface groundwater mix, are recognized as hotspots for biogeochemical reactions; however, little is known of the microbial communities that control many of those reactions. This study investigated the potential functions of microbes inhabiting a cenote and an offshore submarine spring (Pargos) in the near-coastal waters of the Yucatan Peninsula, Mexico. The inland cenote (Cenote Siete Bocas; C7B) is characterized by a chemocline that is host to an array of physicochemical gradients associated with microbial activities. The chemocline includes an increasing gradient in sulfide concentrations with depth and a decreasing gradient in nitrate concentrations. The microbial community within the chemocline was dominated by Sulfurimonas and Sulfurovum of the Campylobacteria, which are likely responsible for sulfide oxidation coupled with nitrate reduction. Although C7B has not been directly connected with Pargos Spring, water discharging from the spring has physicochemical characteristics and microbial community structures similar to C7B, strongly suggesting biogeochemical processing in the STE impacts groundwater composition prior to discharge. This work yields insight into the microbial communities and biogeochemical reactions in STEs in karstic aquifers and provides evidence for the importance of Campylobacteria in controlling nitrate concentrations exported to marine springs.     

Metabolic engineering of the anaerobic central metabolic pathway in Escherichia coli for the simultaneous anaerobic production of isoamyl acetate and succinic acid

An in vivo method of producing isoamyl acetate and succinate simultaneously has been developed in Escherichia coli to maximize yields of both high value compounds as well as maintain the proper redox balance between NADH and NAD⁺. Previous attempts at producing the ester isoamyl acetate anaerobically did not produce the compound in high concentrations because of competing pathways and the need for NAD⁺ regeneration. The objective of this study is to produce succinate as an example of a reduced coproduct to balance the ratio of NADH/NAD⁺ as a way of maximizing isoamyl acetate production. Because the volatility of the two compounds differs greatly, the two could be easily separated in an industrial setting. An ldhA, adhE double mutant strain (SBS110MG) served as the control strain to test the effect of an additional ackA‐pta mutation as found in SBS990MG. Both strains overexpressed the two heterologous genes pyruvate carboxylase and alcohol acetyltransferase (for ester production). The triple mutant SBS990MG was found to produce higher levels of both isoamyl acetate and succinate. At the optimal condition of 25°C, the culture produced 9.4 mM isoamyl acetate and 45.5 mM succinate. SBS990MG produced 36% more ester and over 700% more succinate than SBS110MG. In addition, this study demonstrated that a significantly higher isoamyl acetate concentration can be attained by simultaneously balancing the carbon and cofactor flow; the isoamyl acetate concentration of 9.4 mM is more than seven times higher than an earlier report of about 1.2 mM. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Molecular- and cultivation-based analyses of microbial communities in oil field water and in microcosms amended with nitrate to control H2S production

Nitrate injection into oil fields is an alternative to biocide addition for controlling sulfide production (‘souring’) caused by sulfate-reducing bacteria (SRB). This study examined the suitability of several cultivation-dependent and cultivation-independent methods to assess potential microbial activities (sulfidogenesis and nitrate reduction) and the impact of nitrate amendment on oil field microbiota. Microcosms containing produced waters from two Western Canadian oil fields exhibited sulfidogenesis that was inhibited by nitrate amendment. Most probable number (MPN) and fluorescent in situ hybridization (FISH) analyses of uncultivated produced waters showed low cell numbers (≤10³ MPN/ml) dominated by SRB (>95% relative abundance). MPN analysis also detected nitrate-reducing sulfide-oxidizing bacteria (NRSOB) and heterotrophic nitrate-reducing bacteria (HNRB) at numbers too low to be detected by FISH or denaturing gradient gel electrophoresis (DGGE). In microcosms containing produced water fortified with sulfate, near-stoichiometric concentrations of sulfide were produced. FISH analyses of the microcosms after 55 days of incubation revealed that Gammaproteobacteria increased from undetectable levels to 5–20% abundance, resulting in a decreased proportion of Deltaproteobacteria (50–60% abundance). DGGE analysis confirmed the presence of Delta- and Gammaproteobacteria and also detected Bacteroidetes. When sulfate-fortified produced waters were amended with nitrate, sulfidogenesis was inhibited and Deltaproteobacteria decreased to levels undetectable by FISH, with a concomitant increase in Gammaproteobacteria from below detection to 50–60% abundance. DGGE analysis of these microcosms yielded sequences of Gamma- and Epsilonproteobacteria related to presumptive HNRB and NRSOB (Halomonas, Marinobacterium, Marinobacter, Pseudomonas and Arcobacter), thus supporting chemical data indicating that nitrate-reducing bacteria out-compete SRB when nitrate is added.     

Influence of the pH and redox potential on phosphate activity in the Parana Medio system

The effect of redox potential and pH on the phosphate mobility in two sediments were investigated using both consolidated and suspended sediments from the area where the Parana Medio long reservoir (Atgentina) is to be built (Smirnov, 1984). In addition to direct chemical sediment analysis, extraction techniques were carried out with a stepwise NH₄Cl-NaOH-HCl shaking method, the latter supposedly separating the weakly bound, the Fe- and Al- bound and the Ca- bound phosphates in the sediments.Phosphate released into water depends upon redox potential and pH, which both were modified in an experimental setup. The source of the phosphate was the fraction of Fe and/or Al bound phosphate present both in the sediment and in the suspended solids.Abbreviations cm centimeter - km kilometer - gg gram - l liter - ¬m micrometer - °C grade centigrades - km² square kilometer - m.s^–1 meter per second - m³.s^–1 cubic meter per second - mg.1¹ miligram per liter     

Effect of hydrological conditions on nitrous oxide, methane, and carbon dioxide dynamics in a bottomland hardwood forest and its implication for soil carbon sequestration

This study was conducted at three locations in a bottomland hardwood forest with a distinct elevation and hydrological gradient: ridge (high, dry), transition, and swamp (low, wet). At each location, concentrations of soil greenhouse gases (N₂O, CH₄, and CO₂), their fluxes to the atmosphere, and soil redox potential (Eh) were measured bimonthly, while the water table was monitored every day. Results show that soil Eh was significantly (P < 0.001) correlated with water table: a negative correlation at the ridge and transition locations, but a positive correlation at the permanently flooded swamp location. Both soil gas profile analysis and surface gas flux measurements indicated that the ridge and transition locations could be a sink of atmospheric CH₄, especially in warm seasons, but generally functioned as a minor source of CH₄ in cool seasons. The swamp location was a major source of CH₄, and the emission rate was higher in the warm seasons (mean 28 and median 23 mg m^?2 h^?1) than in the cool seasons (both mean and median 13 mg m^?2 h^?1). Average CO₂ emission rate was 251, 380 and 52 mg m^?2 h^?1 for the ridge, transition and swamp location, respectively. At each location, higher CO₂ emission rates were also found in the warm seasons. The lowest CO₂ emission rate was found at the swamp location, where soil C content was the highest, due to less microbial biomass, less CO₂ production in such an anaerobic environment, and greater difficulty of CO₂ diffusion to the atmosphere. Cumulative global warming potential emission from these three greenhouse gases was in an order of swamp > transition > ridge location. The ratio CO₂/CH₄ production in soil is a critical factor for evaluating the overall benefit of soil C sequestration, which can be greatly offset by CH₄ production and emission.     

Assessing the impact of rumen microbial communities on methane emissions and production traits in Holstein cows in a tropical climate

The evaluation of how the gut microbiota affects both methane emissions and animal production is necessary in order to achieve methane mitigation without production losses. Toward this goal, the aim of this study was to correlate the rumen microbial communities (bacteria, archaea, and fungi) of high (HP), medium (MP), and low milk producing (LP), as well as dry (DC), Holstein dairy cows in an actual tropical production system with methane emissions and animal production traits. Overall, DC cows emitted more methane, followed by MP, HP and LP cows, although HP and LP cow emissions were similar. Using next-generation sequencing, it was found that bacteria affiliated with Christensenellaceae, Mogibacteriaceae, S24-7, Butyrivibrio, Schwartzia, and Treponema were negatively correlated with methane emissions and showed positive correlations with digestible dry matter intake (dDMI) and digestible organic matter intake (dOMI). Similar findings were observed for archaea in the genus Methanosphaera. The bacterial groups Coriobacteriaceae, RFP12, and Clostridium were negatively correlated with methane, but did not correlate with dDMI and dOMI. For anaerobic fungal communities, no significant correlations with methane or animal production traits were found. Based on these findings, it is suggested that manipulation of the abundances of these microbial taxa may be useful for modulating methane emissions without negatively affecting animal production.     

Redox chemistry and chemical biology of H2S,hydropersulfides, and derived species: Implications of their possible biological activity and utility

Hydrogen sulfide (H₂S) is an endogenously generated and putative signaling/effector molecule. Despite its numerous reported functions, the chemistry by which it elicits its functions is not understood. Moreover, recent studies allude to the existence of other sulfur species besides H₂S that may play critical physiological roles. Herein, the basic chemical biology of H₂S as well as other related or derived species is discussed and reviewed. This review particularly focuses on the per- and polysulfides which are likely in equilibrium with free H₂S and which may be important biological effectors themselves.     

Evaluating rRNA as an indicator of microbial activity in environmental communities: limitations and uses

Microbes exist in a range of metabolic states (for example, dormant, active and growing) and analysis of ribosomal RNA (rRNA) is frequently employed to identify the ‘active'' fraction of microbes in environmental samples. While rRNA analyses are no longer commonly used to quantify a population''s growth rate in mixed communities, due to rRNA concentration not scaling linearly with growth rate uniformly across taxa, rRNA analyses are still frequently used toward the more conservative goal of identifying populations that are currently active in a mixed community. Yet, evidence indicates that the general use of rRNA as a reliable indicator of metabolic state in microbial assemblages has serious limitations. This report highlights the complex and often contradictory relationships between rRNA, growth and activity. Potential mechanisms for confounding rRNA patterns are discussed, including differences in life histories, life strategies and non-growth activities. Ways in which rRNA data can be used for useful characterization of microbial assemblages are presented, along with questions to be addressed in future studies.     

Regulation of the activity of glyceraldehyde 3-phosphate dehydrogenase by glutathione and H2O2

The activity lost during storage of a solution of muscle glyceraldehyde 3-phosphate dehydrogenase was rapidly restored on adding a thiol compound, but not arsenite or azide. On treatment with H₂O₂, the enzyme was partially inactivated and complete loss of activity occurred in the presence of glutathione. Samples of the enzyme pretreated with glutathione followed by removal of the thiol compound by filtration on a Sephadex column showed both full activity and its complete loss on adding H₂O₂, in the absence of added glutathione. Most of the activity was restored when the H₂O₂-inactivated enzyme was incubated with glutathione (25mM) or dithiothreitol (5mM) whereas arsenite or azide were partly effective and ascorbate was ineffective. The need for incubation for a long time with a strong reducing agent for restoration of activity suggests that the oxidized group (disulfide or sulfenate) must be in a masked state in the H₂O₂-inactivated enzyme. Analysis by SDS-PAGE gave evidence for the formation of a small quantity of glutathione-reversible disulfide-form of the enzyme. Circular dichroic spectra indicated a decrease in -helical content in the inactivated form of the enzyme. The evidence suggest that glutathione and H₂O₂ can regulate the active state of this enzyme.