Experimental Incubations Elicit Profound Changes in Community Transcription in OMZ Bacterioplankton

Sequencing of microbial community RNA (metatranscriptome) is a useful approach for assessing gene expression in microorganisms from the natural environment. This method has revealed transcriptional patterns in situ, but can also be used to detect transcriptional cascades in microcosms following experimental perturbation. Unambiguously identifying differential transcription between control and experimental treatments requires constraining effects that are simply due to sampling and bottle enclosure. These effects remain largely uncharacterized for “challenging” microbial samples, such as those from anoxic regions that require special handling to maintain in situ conditions. Here, we demonstrate substantial changes in microbial transcription induced by sample collection and incubation in experimental bioreactors. Microbial communities were sampled from the water column of a marine oxygen minimum zone by a pump system that introduced minimal oxygen contamination and subsequently incubated in bioreactors under near in situ oxygen and temperature conditions. Relative to the source water, experimental samples became dominated by transcripts suggestive of cell stress, including chaperone, protease, and RNA degradation genes from diverse taxa, with strong representation from SAR11-like alphaproteobacteria. In tandem, transcripts matching facultative anaerobic gammaproteobacteria of the Alteromonadales (e.g., Colwellia) increased 4–13 fold up to 43% of coding transcripts, and encoded a diverse gene set suggestive of protein synthesis and cell growth. We interpret these patterns as taxon-specific responses to combined environmental changes in the bioreactors, including shifts in substrate or oxygen availability, and minor temperature and pressure changes during sampling with the pump system. Whether such changes confound analysis of transcriptional patterns may vary based on the design of the experiment, the taxonomic composition of the source community, and on the metabolic linkages between community members. These data highlight the impressive capacity for transcriptional changes within complex microbial communities, underscoring the need for caution when inferring in situ metabolism based on transcript abundances in experimental incubations

Microelectrode Studies of Interstitial Water Chemistry and Photosynthetic Activity in a Hot Spring Microbial Mat

Microelectrodes were used to measure oxygen, pH, and oxygenic photosynthetic activity in a hot spring microbial mat (Octopus Spring, Yellowstone National Park), where the cyanobacterium Synechococcus lividus and the filamentous bacterium Chloroflexus aurantiacus are the only known phototrophs. The data showed very high biological activities in the topmost layers of the microbial mat, resulting in extreme values for oxygen and pH. At a 1-mm depth at a 55°C site, oxygen and pH reached 900 μM and 9.4, respectively, just after solar noon, whereas anoxic conditions with a pH of 7.2 were measured before sunrise. Although diurnal changes between these extremes occurred over hours during a diurnal cycle, microbial activity was great enough to give the same response in 1 to 2 min after artificial shading. Oxygenic photosynthesis was confined to a 0.5- to 1.1-mm layer at sites with temperatures at or above about 50°C, with maximum activities in the 55 to 60°C region. The data suggest that S. lividus is the dominant primary producer of the mat

Hydrogen Dynamics in Cyanobacteria Dominated Microbial Mats Measured by Novel Combined H2/H2S and H2/O2 Microsensors

Hydrogen may accumulate to micromolar concentrations in cyanobacterial mat communities from various environments, but the governing factors for this accumulation are poorly described. We used newly developed sensors allowing for simultaneous measurement of H2S and H2 or O2 and H2 within the same point to elucidate the interactions between oxygen, sulfate reducing bacteria, and H2 producing microbes. After onset of darkness and subsequent change from oxic to anoxic conditions within the uppermost ∼1 mm of the mat, H2 accumulated to concentrations of up to 40 μmol L-1 in the formerly oxic layer, but with high variability among sites and sampling dates. The immediate onset of H2 production after darkening points to fermentation as the main H2 producing process in this mat. The measured profiles indicate that a gradual disappearance of the H2 peak was mainly due to the activity of sulfate reducing bacteria that invaded the formerly oxic surface layer from below, or persisted in an inactive state in the oxic mat during illumination. The absence of significant H2 consumption in the formerly oxic mat during the first ∼30 min after onset of anoxic conditions indicated absence of active sulfate reducers in this layer during the oxic period. Addition of the methanogenesis inhibitor BES led to increase in H2, indicating that methanogens contributed to the consumption of H2. Both H2 formation and consumption seemed unaffected by the presence/absence of H2S

CO₂ and O₂ dynamics in leaves of aquatic plants with C₃ or CAM photosynthesis - application of a novel CO₂ microsensor

Background and Aims Leaf tissue CO2 partial pressure (pCO2) shows contrasting dynamics over a diurnal cycle in C3 and Crassulacean Acid Metabolism (CAM) plants. However, simultaneous and continuous monitoring of pCO2 and pO2 in C3 and CAM plants under the same conditions was lacking. Our aim was to use a new CO2 microsensor and an existing O2 microsensor for non-destructive measurements of leaf pCO2 and pO2 dynamics to compare a C3 and a CAM plant in an aquatic environment. Methods A new amperometric CO2 microsensor and an O2 microsensor elucidated with high temporal resolution the dynamics in leaf pCO2 and pO2 during light-dark cycles for C3 Lobelia dortmanna and CAM Littorella uniflora aquatic plants. Underwater photosynthesis, dark respiration, tissue malate concentrations and sediment CO2 and O2 were also measured. Key Results During the dark period, for the C3 plant, pCO2 increased to approx. 3.5 kPa, whereas for the CAM plant CO2 was mostly below 0.05 kPa owing to CO2 sequestration into malate. Upon darkness, the CAM plant had an initial peak in pCO2 (approx. 0.16 kPa) which then declined to a quasi-steady state for several hours and then pCO2 increased towards the end of the dark period. The C3 plant became severely hypoxic late in the dark period, whereas the CAM plant with greater cuticle permeability did not. Upon illumination, leaf pCO2 declined and pO2 increased, although aspects of these dynamics also differed between the two plants. Conclusions The continuous measurements of pCO2 and pO2 highlighted the contrasting tissue gas compositions in submerged C3 and CAM plants. The CAM leaf pCO2 dynamics indicate an initial lag in CO2 sequestration to malate, which after several hours of malate synthesis then slows. Like the use of O2 microsensors to resolve questions related to plant aeration, deployment of the new CO2 microsensor will benefit plant ecophysiology research