Effect of titanium dioxide concentration on the survival of Pseudomonas stutzeri during irradiation with near ultraviolet light

Cells of Pseudomonas stutzeri in suspensions of TiO₂ ranging in concentration from 0.5 to 4.0 g l^-1 were irradiated with a blacklight blue u.v. source displaying peak emissivity at approximately 370 mm. Irradiation under these conditions is known to result in the generation of lethal free radicals. During irradiation the suspensions were agitated, using a specially modified laboratory shaker, to ensure efficient exposure of the TiO₂. A u.v. radiation dose of 175 kJ m^-2 resulted in cell fractional survival ranging from 5.5 times 10^-5, at the lowest TiO₂ concentration, to 1.0 times 10^-6, at the highest TiO₂ concentration. The advantages of contactors employing TiO₂ suspensions are briefly compared to immobilized TiO₂ systems.     

Photoelectrochemical sterilization of microbial cells by semiconductor powders

Abstract We report the novel concept of photochemical sterilization. Microbial cells were killed photoelectrochemically with semiconductor powder (platinum-loaded titanium oxide, TiO₂/Pt). Coenzyme A, (CoA) in the whole cells was photo-electrochemically oxidized and, as a result, the respiration of cells was inhibited. Inhibition of respiratory activity caused death of the cells. Lactobacillus acidophilus, Saccharomyces cerevisiae and Escherichia coli (10³ cells/ml respectively) were completely sterilized when they were incubated with TiO₂/Pt particles under metal halide lamp irradiation for 60–120 min.     

Evacuation of pelleted feed and the suitability of titanium(IV) oxide as a feed marker for gut kinetics in Nile tilapia

The present study assessed the suitability of titanium(IV) oxide, TiO₂, as a digesta passage marker in Nile tilapia Oreochromis niloticus and studied the shape of the evacuation curve in this species. In three separate trials, fish were given one dose of either 0·5, 0·25 or 0·1% of their body mass (% BME) of feed marked with 1% TiO₂ or 0·5% BME of the same feed without marker. The fish were serially slaughtered at intervals after feeding and the stomach contents analysed for dry mass and marker content. The data for individual trials were analysed with the linear, square root, surface area and exponential evacuation models and parameter comparisons showed that, although the marker interfered slightly with the evacuation process, true meal size could be predicted more accurately from the marker data. The results of an analysis of the combined data sets suggested that stomach evacuation in this species is dependent more on food particle surface area (surface area model) than on stomach content mass (exponential model) as is generally assumed. On the basis of these results, it was concluded that TiO₂ at an inclusion level of 1% is an acceptable marker for quantifying evacuation with a view to predicting food consumption but should be used with caution in digestibility studies.     

Soil microbial responses to increased concentrations of atmospheric CO2

Terrestrial ecosystems respond to an increased concentration of atmospheric CO₂. While elevated atmospheric CO₂ has been shown to alter plant growth and productivity, it also affects ecosystem structure and function by changing below-ground processes. Knowledge of how soil microbiota respond to elevated atmospheric CO₂ is of paramount importance for understanding global carbon and nutrient cycling and for predicting changes at the ecosystem-level. An increase in the atmospheric CO₂ concentration not only alters the weight, length, and architecture of plant roots, but also affects the biotic and abiotic environment of the root system. Since the concentration of CO₂ in soil is already 10–50 times higher than that in the atmosphere, it is unlikely that increasing atmospheric CO₂ will directly influence the rhizosphere. Rather, it is more likely that elevated atmospheric CO₂ will affect the microbe–soil–plant root system indirectly by increasing root growth and rhizodeposition rates, and decreasing soil water deficit. Consequently, the increased amounts and altered composition of rhizosphere-released materials will have the potential to alter both population and community structure, and activity of soil- and rhizosphere-associated microorganisms. This occurrence could in turn affect plant health and productivity and plant community structure. This review covers current knowledge about the response of soil microbes to elevated concentrations of atmospheric CO₂.     

Stimulation of r- vs. K-selected microorganisms by elevated atmospheric CO2 depends on soil aggregate size

Increased root exudation under elevated atmospheric CO₂ and the contrasting environments in soil macro- and microaggregates could affect microbial growth strategies. We investigated the effect of elevated CO₂ on the contribution of fast- ( r -strategists) and slow-growing ( K -strategists) microorganisms in soil macro- and microaggregates. We fractionated the bulk soil from the ambient and elevated (for 5 years) CO₂ treatments of FACE-Hohenheim (Stuttgart) into large macro- (>2 mm), small macro- (0.25–2.00 mm), and microaggregates (<0.25 mm) using 'optimal moist' sieving. Microbial biomass (C_mic), the maximum specific growth rate (μ), growing microbial biomass (GMB) and lag-period ( t _lag) were estimated by the kinetics of CO₂ emission from bulk soil and aggregates amended with glucose and nutrients. Although C_org and C_mic were unaffected by elevated CO₂, μ values were significantly higher under elevated than ambient CO₂ for bulk soil, small macroaggregates, and microaggregates. Substrate-induced respiratory response increased with decreasing aggregate size under both CO₂ treatments. Based on changes in μ, GMB and lag period, we conclude that elevated atmospheric CO₂ stimulated the r- selected microorganisms, especially in soil microaggregates. Such an increase in r -selected microorganisms indicates acceleration of available C mineralization in soil, which may counterbalance the additional C input by roots in soils in a future elevated atmospheric CO₂ environment.     

Elevated atmospheric CO2 affects soil microbial diversity associated with trembling aspen

The effects of elevated atmospheric CO₂ (560 p.p.m.) and subsequent plant responses on the soil microbial community composition associated with trembling aspen was assessed through the classification of 6996 complete ribosomal DNA sequences amplified from the Rhinelander WI free-air CO₂ and O₃ enrichment (FACE) experiments microbial community metagenome. This in-depth comparative analysis provides an unprecedented, detailed and deep branching profile of population changes incurred as a response to this environmental perturbation. Total bacterial and eukaryotic abundance does not change; however, an increase in heterotrophic decomposers and ectomycorrhizal fungi is observed. Nitrate reducers of the domain bacteria and archaea, of the phylum Crenarchaea , potentially implicated in ammonium oxidation, significantly decreased with elevated CO₂. These changes in soil biota are evidence for altered interactions between trembling aspen and the microorganisms in its surrounding soil, and support the theory that greater plant detritus production under elevated CO₂ significantly alters soil microbial community composition.     

Mhp493 (P216) is a proteolytically processed, cilium and heparin binding protein of Mycoplasma hyopneumoniae

Mycoplasma hyopneumoniae induces respiratory disease in swine by colonizing cilia causing ciliostasis, cilial loss and epithelial cell death. Heparin binds to M. hyopneumoniae cells in a dose-dependent manner and blocks its ability to adhere to porcine cilia. We show here that Mhp493 (P216), a paralogue of the cilium adhesin P97 (Mhp183), is cleaved between amino acids 1040 and 1089 generating surface-accessible, heparin-binding proteins P120 and P85. Antiphosphoserine antibodies recognized P85 in 2-D immunoblotting studies and TiO₂ chromatography of trypsin digests of P85 isolated a single peptide with an m/z of 917.3. A phosphoserine residue in the tryptic peptide ⁹⁰VSELpSFR⁹⁶ (position 94 in P85) was identified by MALDI-MS/MS. Polyhistidine fusion proteins (F1_P216, F2_P216, F3_P216) spanning Mhp493 bound heparin with biologically significant Kd values, and heparin, fucoidan and mucin inhibited this interaction. Latex beads coated with F1_P216, F2_P216 and F3_P216 adhered to and entered porcine kidney epithelial-like (PK15) cell monolayers. Microtitre plate-based assays showed that sequences within P120 and P85 bind to porcine cilia and are recognized by serum antibodies elicited during infection by M. hyopneumoniae . Mhp493 contributes significantly to the surface architecture of M. hyopneumoniae and is the first cilium adhesin to be described that lacks an R1 cilium-binding domain.     

Monitoring the fate of genetically engineered bacteria sprayed on the phylloplane of bush beans and grass

Abstract The fate of a Bacillus amyloliquefaciens with the recombinant plasmid pSB20 sprayed on the phyllosphere of grass, and of a Tn 5 marked Pseudomonas syringae sprayed on the phyllosphere of bush beans was studied in planted soil microcosms. B. amyloliquefaciens showed a decline from 1.5×10⁸ to 3.1×10² cfu g⁻¹ on the phylloplane of grass in the course of the experiment. B. amyloliquefaciens was easy to follow by selective cultivation due to the complete absence of bacterial background growth. Southern blot hybridization of Hin dIII digested genomic DNA showed plasmid restriction patterns identical with pSB20 indicating high plasmid stability. In total DNA extracts from phyllosphere bacteria the recombinant plasmid was detectable by Southern blot hybridization up to 6×10⁴ cfu g⁻¹ (wet weight). Counts of hybridizing colonies showed that P. syringae established on the phyllosphere of bush beans at between 5×10³ and 4×10⁶ cfu g⁻¹ fresh weight. During senescence of the bean plants the strain was no longer detectable by selective cultivation and subsequent colony hybridization. In contrast, Tn5 marked DNA was detected after PCR amplification over the whole period of the experiment.     

Links between methanotroph community composition and CH4 oxidation in a pine forest soil

The main gap in our knowledge about what determines the rate of CH₄ oxidation in forest soils is the biology of the microorganisms involved, the identity of which remains unclear. In this study, we used stable-isotope probing (SIP) following ¹³CH₄ incorporation into phospholipid fatty acids (PLFAs) and DNA/RNA, and sequencing of methane mono-oxygenase ( pmoA ) genes, to identify the influence of variation in community composition on CH₄ oxidation rates. The rates of ¹³C incorporation into PLFAs differed between horizons, with low ¹³C incorporation in the organic soil and relatively high ¹³C incorporation into the two mineral horizons. The microbial community composition of the methanotrophs incorporating the ¹³C label also differed between horizons, and statistical analyses suggested that the methanotroph community composition was a major cause of variation in CH₄ oxidation rates. Both PLFA and pmoA -based data indicated that CH₄ oxidizers in this soil belong to the uncultivated 'upland soil cluster α'. CH₄ oxidation potential exhibited the opposite pattern to ¹³C incorporation, suggesting that CH₄ oxidation potential assays may correlate poorly with in situ oxidation rates. The DNA/RNA-SIP assay was not successful, most likely due to insufficient ¹³C-incorporation into DNA/RNA. The limitations of the technique are briefly discussed.     

Anaerobic sulfur oxidation in the absence of nitrate dominates microbial chemoautotrophy beneath the pelagic chemocline of the eastern Gotland Basin, Baltic Sea

Oxic–anoxic interfaces harbor significant numbers and activity of chemolithoautotrophic microorganisms, known to oxidize reduced sulfur or nitrogen species. However, measurements of in situ distribution of bulk carbon dioxide (CO₂) assimilation rates and active autotrophic microorganisms have challenged the common concept that aerobic and denitrifying sulfur oxidizers are the predominant autotrophs in pelagic oxic–anoxic interfaces. Here, we provide a comparative investigation of nutrient, sulfur, and manganese chemistry, microbial biomass distribution, as well as CO₂ fixation at the pelagic redoxcline of the eastern Gotland Basin, Baltic Sea. Opposing gradients of oxygen, nitrate, and sulfide approached the detection limits at the chemocline at 204 m water depth. No overlap of oxygen or nitrate with sulfide was observed, whereas particulate manganese was detected down to 220 m. More than 70% of the bulk dark CO₂ assimilation, totaling 9.3 mmol C m⁻² day⁻¹, was found in the absence of oxygen, nitrite, and nitrate and could not be stimulated by their addition. Maximum fixation rates of up to 1.1 μmol C L⁻¹ day⁻¹ were surprisingly susceptible to altered redox potential or sulfide concentration. These results suggest that novel redox-sensitive pathways of microbial sulfide oxidation could account for a significant fraction of chemolithoautotrophic growth beneath pelagic chemoclines. A mechanism of coupled activity of sulfur-oxidizing and sulfur-reducing microorganisms is proposed.     

Coexistence of nitrifiers, denitrifiers and Anammox bacteria in a sequencing batch biofilm reactor as revealed by PCR-DGGE

Aims:  The bacterial diversity in a sequencing batch biofilm reactor (SBBR) treating landfill leachate was studied to explain the mechanism of nitrogen removal.
Methods and Results:  The total microbial DNA was extracted from samples collected from landfill leachate and biofilm of the reactor with the removal efficiencies of NH₄⁺-N higher than 97% and that of chemical oxygen demand (determined by K₂Cr₂O₇, COD_Cr) higher than 86%. Denaturing gradient gel electrophoresis (DGGE) fingerprints based on total community 16S rRNA genes were analyzed with statistical methods, and excised DNA bands were sequenced. The results of phylogenetic analyses revealed high diversity within the SBBR biofilm community, and DGGE banding patterns showed that the community structure in the biofilm remained stable during the running period.
Conclusions:  A coexistence of nitrifiers, including ammonia-oxidizing bacteria and nitrite-oxidizing bacteria, denitrifiers, including aerobic or anaerobic denitrifying bacteria and Anammox bacteria were detected, which might be the real matter of high removal efficiencies of NH₄⁺-N and COD_Cr in the reactor.
Significance and Impact of the Study:  The findings in this study indicated that PCR-DGGE analysis could be used for microbial community detection as prior method, and the SBBR technique could provide preferable growing environment for bacteria with N removal function.     

Inorganic carbon limitation of photosynthesis in lake phytoplankton

1. Inorganic carbon availability influences species composition of phytoplankton in acidic and highly alkaline lakes, whereas the overall influence on community photosynthesis and growth is subject to debate.
2. The influence of total dissolved inorganic carbon (DIC) and free CO₂ on community photosynthesis was studied in six Danish lakes during the summer of 1995. The lakes were selected to ensure a wide range of chlorophyll a concentrations (1–120 μg l^–1), pH (5.6–9.6) and DIC concentration (0.02–2.5 m m ). Photosynthesis experiments were performed using the ¹⁴C technique in CO₂-manipulated water samples, either by changing the pH or by adding/removing CO₂.
3. Lake waters were naturally CO₂ supersaturated during most of the experimental period and inorganic carbon limitation of photosynthetic rates did not occur under ambient conditions. However, photosynthesis by phytoplankton in lakes with low and intermediate DIC concentrations was seriously restricted when CO₂ concentrations declined. Similarly, photosynthesis was limited by low CO₂ concentrations during phytoplankton blooms in the hardwater alkaline lakes.     

Intraspecific variation in the response of rice (Oryza sativa) to increased CO2– photosynthetic, biomass and reproductive characteristics

Two rice ( Oryza sativa L.) cultivars of contrasting morphologies, IR-36 and Fujiyama-5, were exposed to ambient (360 μl l⁻¹) and ambient plus 300 μl l⁻¹ CO₂ from time of emergence until ca 50% grain fill at the Duke University Phytotron, Durham, North Carolina. Exposure to increased CO₂ resulted in about a 50% increase in the photosynthetic rate for both cultivars and photosynthetic enhancement was still evident after 3 months of exposure to a high CO₂ environment. The photosynthetic response at 5% CO₂ and the response of CO₂ assimilation (A) to internal CO₂ (C_i) suggest a reallocation of biochemical resources from RuBP carboxylation to RuBP regeneration. Increases in total plant biomass at elevated CO₂ were approximately the same in both cultivars, although differences in allocation patterns were noted in root/shoot ratio. Differences in reproductive characteristics were also observed between cultivars at an elevated CO₂ environment with a significant increase in harvest index for IR-36 but not for Fujiyama-5. Changes in carbon allocation in reproduction between these two cultivars suggest that lines of rice could be identified that would maximize reproductive output in a future high CO₂ environment.     

Colloidal suspensions of clay or titanium dioxide nanoparticles can inhibit leaf growth and transpiration via physical effects on root water transport

A laboratory investigation was conducted to determine whether colloidal suspensions of inorganic nanoparticulate materials of natural or industrial origin in the external water supplied to the primary root of maize seedlings ( Zea mays L.) could interfere with water transport and induce associated leaf responses. Water flow through excised roots was reduced, together with root hydraulic conductivity, within minutes of exposure to colloidal suspensions of naturally derived bentonite clay or industrially produced TiO₂ nanoparticles. Similar nanoparticle additions to the hydroponic solution surrounding the primary root of intact seedlings rapidly inhibited leaf growth and transpiration. The reduced water availability caused by external nanoparticles and the associated leaf responses appeared to involve a rapid physical inhibition of apoplastic flow through nanosized root cell wall pores rather than toxic effects. Thus: (1) bentonite and TiO₂ treatments also reduced the hydraulic conductivity of cell wall ghosts of killed roots left after hot alcohol disruption of the cell membranes; and (2) the average particle exclusion diameter of root cell wall pores was reduced from 6.6 to 3.0 nm by prior nanoparticle treatments. Irrigation of soil-grown plants with nanoparticle suspensions had mostly insignificant inhibitory effects on long-term shoot production, and a possible developmental adaptation is suggested.     

13C-isotope analyses reveal that chemolithoautotrophic Gamma- and Epsilonproteobacteria feed a microbial food web in a pelagic redoxcline of the central Baltic Sea

Marine pelagic redoxclines are zones of high dark CO₂ fixation rates, which can correspond up to 30% of the surface primary production. However, despite this significant contribution to the pelagic carbon cycle, the identity of most chemolithoautotrophic organisms is still unknown. Therefore, the aim of this study was to directly link the dark CO₂ fixation capacity of a pelagic redoxcline in the central Baltic Sea (Landsort Deep) with the identity of the main chemolithoautotrophs involved. Our approach was based on the analysis of natural carbon isotope signatures in fatty acid methyl esters (FAMEs) and on measurements of CO₂ incorporation in ¹³C-bicarbonate pulse experiments. The incorporation of ¹³C into chemolithoautotrophic cells was investigated by rRNA-based stable isotope probing (RNA-SIP) and FAME analysis after incubation for 24 and 72 h under in situ conditions. Our results demonstrated that fatty acids indicative of Proteobacteria were significantly enriched in ¹³C slightly below the chemocline. RNA-SIP analyses revealed that two different Gammaproteobacteria and three closely related Epsilonproteobacteria of the Sulfurimonas cluster were active dark CO₂-fixing microorganisms, with a time-dependent community shift between these groups. Labelling of Archaea was not detectable, but after 72 h of incubation the ¹³C-label had been transferred to a potentially bacterivorous ciliate related to Euplotes sp. Thus, RNA-SIP provided direct evidence for the contribution of chemolithoautotrophic production to the microbial food web in this marine pelagic redoxcline, emphasizing the importance of dark CO₂-fixing Proteobacteria within this habitat.     

Effects of phenolic acids on ammonia oxidation by terrestrial autotrophic nitrifying microorganisms

Abstract It has been hypothesized that vegetation in certain ecosystems inhibits nitrification in soil by producing phenolic compounds that inhibit oxidation of ammonia by nitrifying microorganisms. This hypothesis is based largely on a report that very low concentrations (10⁻⁶ M–10⁻⁸ M) of several phenolic acids (notably ferulic acid) completely inhibited NO₂⁻ production in an aqueous suspension of soil treated with (NH₄)₂SO₄ and a nutrient solution suitable for growth of Nitrosomonas and other autotrophic nitrifying microorganisms. To evaluate this hypothesis, we determined the effects of three ohenolic acids (ferulic acid, caffeic acid, and p -coumaric on nitrite production by representatives of three genera of terrestrial autotrophic nitrifying microorganisms ( Nitrosospira, Nitrosomonas , or Nitrosolubos ) grown on a defined medium containing NH₄⁺. We found that nitrite production by the Nitrososspira was not inhibited by ferulic acid, caffeic acid, or p -coumaric acid at concentrations of 10⁻⁶ or 10⁻⁵ M and was only slightly inhibited when these acids were at a concentration of 10⁻⁴ M. We also found that ferulic acid did not markedly inhibit nitrite production by the three genera of nitrifying microorganisms studied, even when its concentration was as high as 10⁻³ M. These observations invalidate the hypothesis tested because the phenolic acids studied did not significantly retard ammonia oxidation by autotrophic microorganisms even when their concentration in cultures of these microorganisms greatly exceeded their concentrations in soils.     

Acetogenesis from H2 and CO2 by methane- and non-methane-producing human colonic bacterial communities

Abstract: The purpose of the study was to define the potential for reductive acetogenesis of colonic microflora from six non-methane- and four methane-excreting human subjects in relation to numbers of the different H₂-utilizing microorganisms. Faecal bacterial suspensions were incubated in the presence of NaH¹³CO₃ and under a gas phase composed of either 100% N₂ (control) or 80% H₂–20% N₂. The effects of a specific methanogenesis inhibitor or of sulfate supplementation were also determined. Quantitative nuclear magnetic resonance showed the presence of both single- and double-labelled acetate in all incubations under hydrogen. H₂/CO₂-acetogenesis appears to be a quantitatively important activity only in the presence of very low numbers of methanogens. Inhibition of methanogenesis induced a large increase in ¹³CO₂ incorporation into acetate in CH₄-producing samples. These results showed that methanogens can efficiently outcompete acetogens in human colonic contents. In contrast, no clear-cut competition for H₂ between acetogenesis and dissimilatory sulfate-reduction could be demonstrated. A slight reduction of the acetogenic activity was only observed at the highest sulfate addition (100 mM).     

Growth in elevated CO2 enhances temperature response of photosynthesis in wheat

The temperature dependence of C₃ photosynthesis may be altered by the growth environment. The effects of long-term growth in elevated CO₂ on photosynthesis temperature response have been investigated in wheat ( Triticum aestivum L.) grown in controlled chambers with 370 or 700 μmol mol⁻¹ CO₂ from sowing through to anthesis. Gas exchange was measured in flag leaves at ear emergence, and the parameters of a biochemical photosynthesis model were determined along with their temperature responses. Elevated CO₂ slightly decreased the CO₂ compensation point and increased the rate of respiration in the light and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) V_cmax, although the latter effect was reversed at 15°C. With elevated CO₂, J_max decreased in the 15–25°C temperature range and increased at 30 and 35°C. The temperature response (activation energy) of V_cmax and J_max increased with growth in elevated CO₂. CO₂ enrichment decreased the ribulose 1,5-bisphosphate (RuBP)-limited photosynthesis rates at lower temperatures and increased Rubisco- and RuBP-limited rates at higher temperatures. The results show that the photosynthesis temperature response is enhanced by growth in elevated CO₂. We conclude that if temperature acclimation and factors such as nutrients or water availability do not modify or negate this enhancement, the effects of future increases in air CO₂ on photosynthetic electron transport and Rubisco kinetics may improve the photosynthetic response of wheat to global warming.     

Relationship of environmental pCO2 to selected blood parameters of Oncorhynchus kisutch (Walbaum)

Blood samples from sub-adult coho salmon ( Oncorhynchus kisutch ) were correlated with environmental conditions. Carbon dioxide tensions, pH and bicarbonate levels were determined in the blood for fish acclimated to environmental pCO₂ levels of 2.49 and 2.74 mmHg. Fish were removed from the high pCO₂ environment and placed in one with a low pCO₂. Their blood pH and pCO₂ were monitored during the course of acclimation. After 27 days of acclimation to a low pCO₂ environment, a group of fish was sampled and compared to a group of fish acclimated to a high pCO₂ environment. It was found that pCO₂ in the environment affects the rate of diffusion of carbon dioxide from the blood to the water. Under the environmental conditions prescribed in these experiments no significant change in bicarbonate was noticed. It is postulated that a bicarbonate chloride exchange is not a major pathway for the removal of carbon dioxide from the blood of O. kisutch .     

Changes in abundance, composition and controls within the plankton of a fertilised arctic lake

1. An oligotrophic arctic lake was fertilised with inorganic nitrogen and phosphorus as (NH₄)₂ NO₃ and H₃PO₄ for five summers. The loading rate was 1.7–2.5 mmol N m^–2 day^–1 and 0.136–0.20 mmol P m^–2 day^–1 which is two to three times the annual loading of lakes in the area. The heterotrophic microzooplankton community was enumerated during the experiment as well as 1 year pre- and post-treatment.
2. The structure of the microplankton community changed from a nutrient limited system, dominated by oligotrich protozoans and small-particle feeding rotifers, to a system dominated by a succession of peritrich protozoans and predatory rotifers. These peritrich protozoans and predatory rotifers were not present prior to fertilisation and never constituted more than a small fraction of the biomass in other lakes at the research site. The average biomass of the rotifers and protozoans was more than seven and a half times larger by the end of fertilisation than it was initially.
3. Because of the increases in numbers of individuals in these new taxa, the structure of the microbial food web changed. When fertilisation stopped, most parameters returned to prefertilisation levels within 1 year.