New generation NMR bioreactor coupled with high-resolution NMR spectroscopy leads to novel discoveries in Moorella thermoacetica metabolic profiles.

An in situ nuclear magnetic resonance (NMR) bioreactor was developed and employed to monitor microbial metabolism under batch growth conditions in real time. We selected Moorella thermoacetica ATCC 49707 as a test case. M.

Biofilm shows spatially stratified metabolic responses to contaminant exposure.

Biofilms are core to a range of biological processes, including the bioremediation of environmental contaminants. Within a biofilm population, cells with diverse genotypes and phenotypes coexist, suggesting that distinct metabolic pathways may be expressed based on the local environmental conditions in a biofilm. However, metabolic responses to local environmental conditions in a metabolically active biofilm interacting with environmental contaminants have never been quantitatively elucidated.

Identifying low pH active and lactate-utilizing taxa within oral microbiome communities from healthy children using stable isotope probing techniques.

Background: Many human microbial infectious diseases including dental caries are polymicrobial in nature. How these complex multi-species communities evolve from a healthy to a diseased state is not well understood. Although many health- or disease-associated oral bacteria have been characterized in vitro, their physiology within the complex oral microbiome is difficult to determine with current approaches.

Contribution of extracellular polymeric substances from Shewanella sp. HRCR-1 biofilms to U(VI) immobilization.

The goal of this study was to quantify the contribution of extracellular polymeric substances (EPS) to U(VI) immobilization by Shewanella sp. HRCR-1. Through comparison of U(VI) immobilization using cells with bound EPS (bEPS) and cells with minimal EPS, we show that (i) bEPS from Shewanella sp.

In situ effective diffusion coefficient profiles in live biofilms using pulsed-field gradient nuclear magnetic resonance.

Diffusive mass transfer in biofilms is characterized by the effective diffusion coefficient. It is well documented that the effective diffusion coefficient can vary by location in a biofilm. The current literature is dominated by effective diffusion coefficient measurements for distinct cell clusters and stratified biofilms showing this spatial variation.

Investigations of structure and metabolism within Shewanella oneidensis MR-1 biofilms.

Biofilms possess spatially and temporally varying metabolite concentration profiles at the macroscopic and microscopic scales. This results in varying growth environments that may ultimately drive species diversity, determine biofilm structure and the spatial distribution of the community members. Using non-invasive nuclear magnetic resonance (NMR) microscopic imaging/spectroscopy and confocal imaging, we investigated the kinetics and stratification of anaerobic metabolism within live biofilms of the dissimilatory metal-reducing bacterium Shewanella oneidensis strain MR-1.

NMR bioreactor development for live in-situ microbial functional analysis.

A live, in-situ metabolomics capability was developed for prokaryotic cultures under controlled growth conditions. Toward this goal, a radiofrequency-transparent bioreactor was developed and integrated with a commercial wide-bore nuclear magnetic resonance (NMR) imaging spectrometer and a commercial bioreactor controller. Water suppressed 1H NMR spectroscopy was used to monitor glucose and fructose utilization and byproduct excretion by Eubacterium aggregans (an anaerobic bacterial species relevant for biofuel production) under controlled batch and continuous culture conditions.

Correlated biofilm imaging, transport and metabolism measurements via combined nuclear magnetic resonance and confocal microscopy.

Bacterial biofilms are complex, three-dimensional communities found nearly everywhere in nature and are also associated with many human diseases. Detailed metabolic information is critical to understand and exploit beneficial biofilms as well as combat antibiotic-resistant, disease-associated forms. However, most current techniques used to measure temporal and spatial metabolite profiles in these delicate structures are invasive or destructive.

Localized in vivo isotropic-anisotropic correlation 1H NMR spectroscopy using ultraslow magic angle spinning.

In a previous work (1), the susceptibility broadening in the (1)H NMR metabolite spectrum obtained in a live mouse was separated from the isotropic information, which significantly increased the spectral resolution. This was achieved using ultraslow magic angle spinning (MAS) of the animal combined with a modified phase-corrected magic angle turning (PHORMAT) pulse sequence. However, PHORMAT cannot be used for spatially selective spectroscopy.

Slow-MAS NMR: A new technology for in vivo metabolomic studies.

Obtaining detailed in vivo metabolic information has been identified as key elements of better understanding the efficacy and toxicity of new therapies. A new nuclear magnetic resonance (NMR) technology called LOCMAT is reported in this paper that yields substantially increased spectral resolution in spatially localized in vivo H NMR metabolite spectra, as illustrated by measurements in the liver of a live mouse. LOCMAT promises to significantly enhance the utility of NMR spectroscopy for biomedical research.

NMR methods for in situ biofilm metabolism studies.

Novel procedures and instrumentation are described for nuclear magnetic resonance (NMR) spectroscopy and imaging studies of live, in situ microbial films. A perfused NMR/optical microscope sample chamber containing a planar biofilm support was integrated into a recirculation/dilution flow loop growth reactor system and used to grow in situ Shewanella oneidensis strain MR-1 biofilms. Localized NMR techniques were developed and used to non-invasively monitor time-resolved metabolite concentrations and to image the biomass volume and distribution.

Simultaneous 1H PFG-NMR and confocal microscopy of monolayer cell cultures: effects of apoptosis and necrosis on water diffusion and compartmentalization.

We induced apoptosis and necrosis in monolayer cultures of Chinese hamster ovary cells using okadaic acid and hydrogen peroxide (H2O2), respectively, and examined the effect on water diffusion and compartmentalization using pulsed-field-gradient (PFG) 1H-NMR and simultaneous confocal microscopy. In PFG experiments characterized by a fixed diffusion time (<4.7 ms) and variable b-values (0-27000 s/mm2), 1H-NMR data collected with untreated cells exhibited multiexponential behavior.