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At least 37 records · Page 2

Low-shear modeled microgravity: a global environmental regulatory signal affecting bacterial gene expression, physiology, and pathogenesis

Bacteria inhabit an impressive variety of ecological niches and must adapt constantly to changing environmental conditions. While numerous environmental signals have been examined for their effect on bacteria, the effects of mechanical forces such as shear stress and gravity have only been investigated to a limited extent. However, several important studies have demonstrated a key role for the environmental signals of low shear and/or microgravity in the regulation of bacterial gene expression, physiology, and pathogenesis [Chem. Rec. 1 (2001) 333; Appl. Microbiol. Biotechnol. 54 (2000) 33; Appl. Environ. Microbiol. 63 (1997) 4090; J. Ind. Microbiol. 18 (1997) 22; Curr. Microbiol. 34(4) (1997) 199; Appl. Microbiol. Biotechnol. 56(3-4) (2001) 384; Infect Immun. 68(6) (2000) 3147; Cell 109(7) (2002) 913; Appl. Environ. Microbiol. 68(11) (2002) 5408; Proc. Natl. Acad. Sci. U. S. A. 99(21) (2002) 13807]. The response of bacteria to these environmental signals, which are similar to those encountered during prokaryotic life cycles, may provide insight into bacterial adaptations to physiologically relevant conditions. This review focuses on the current and potential future research trends aimed at understanding the effect of the mechanical forces of low shear and microgravity analogues on different bacterial parameters. In addition, this review also discusses the use of microgravity technology to generate physiologically relevant human tissue models for research in bacterial pathogenesis.

NASA Discipline Cell Biology↗

Rapid in situ assessment of physiological activities in bacterial biofilms using fluorescent probes

Two rapid in situ enumeration methods using fluorescent probes were used to assess the physiological activities of Klebsiella pneumoniae biofilms on stainless steel. Fluorescent dyes, 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and rhodamine 123 (Rh 123), were chosen to perform this study. CTC is a soluble redox indicator which can be reduced by respiring bacteria to fluorescent CTC-formazan crystals. Rh 123 is incorporated into bacteria with respect to cellular proton motive force. The intracellular accumulation of these fluorescent dyes can be determined using epifluorescence microscopy. The results obtained with these two fluorescent probes in situ were compared to the plate count (PC) and in situ direct viable count (DVC) methods. Viable cell densities within biofilms determined by the three in situ methods were comparable and always showed approximately 2-fold higher values than those obtained with the PC method. As an additional advantage, the results were observed after 2 h, which was shorter than the 4 h incubation time required for the DVC method and 24 h for colony formation. The results indicate that staining with CTC and Rh 123 provides rapid information regarding cell numbers and physiological activities of bacteria within biofilms.

NASA Discipline Number 04-10↗

Improved methods for counting bacteria in physiological fluids

Bacterial population detection is based on detection of adenosine triphosphate (ATP), chemical present in all living matter. Amount of ATP in sample, after chemically removing all nonbacterial ATP, is directly related to bacterial population. Sensitivity is improved by concentration step; specificity is improved by lowering pH of solution.

Picciolo, G. L.↗

A natural view of microbial biodiversity within hot spring cyanobacterial mat communities

This review summarizes a decade of research in which we have used molecular methods, in conjunction with more traditional approaches, to study hot spring cyanobacterial mats as models for understanding principles of microbial community ecology. Molecular methods reveal that the composition of these communities is grossly oversimplified by microscopic and cultivation methods. For example, none of 31 unique 16S rRNA sequences detected in the Octopus Spring mat, Yellowstone National Park, matches that of any prokaryote previously cultivated from geothermal systems; 11 are contributed by genetically diverse cyanobacteria, even though a single cyanobacterial species was suspected based on morphologic and culture analysis. By studying the basis for the incongruity between culture and molecular samplings of community composition, we are beginning to cultivate isolates whose 16S rRNA sequences are readily detected. By placing the genetic diversity detected in context with the well-defined natural environmental gradients typical of hot spring mat systems, the relationship between gene and species diversity is clarified and ecological patterns of species occurrence emerge. By combining these ecological patterns with the evolutionary patterns inherently revealed by phylogenetic analysis of gene sequence data, we find that it may be possible to understand microbial biodiversity within these systems by using principles similar to those developed by evolutionary ecologists to understand biodiversity of larger species. We hope that such an approach guides microbial ecologists to a more realistic and predictive understanding of microbial species occurrence and responsiveness in both natural and disturbed habitats.

Review↗

Forster energy transfer in chlorosomes of green photosynthetic bacteria

Energy transfer properties of whole cells and chlorosome antenna complexes isolated from the green sulfur bacteria Chlorobium limicola (containing bacteriochlorophyll c), Chlorobium vibrioforme (containing bacteriochlorophyll d) and Pelodictyon phaeoclathratiforme (containing bacteriochlorophyll e) were measured. The spectral overlap of the major chlorosome pigment (bacteriochlorophyll c, d or, e) with the bacteriochlorophyll a B795 chlorosome baseplate pigment is greatest for bacteriochlorophyll c and smallest for bacteriochlorophyll e. The absorbance and fluorescence spectra of isolated chlorosomes were measured, fitted to gaussian curves and the overlap factors with B795 calculated. Energy transfer times from the bacteriochlorophyll c, d or e to B795 were measured in whole cells and the results interpreted in terms of the Forster theory of energy transfer.

NASA Program Exobiology↗

Physiological assessment of bacteria using fluorochromes

This minireview focuses on the application of fluorogenic compounds in the detection of bacteria with particular emphasis on the assessment of physiological activity using epifluorescence microscopy. Microbiological applications of several related methods will also be reviewed.

NASA Discipline Environmental Health↗

Rhodoferax antarcticus sp. nov., a moderately psychrophilic purple nonsulfur bacterium isolated from an Antarctic microbial mat

A new species of purple nonsulfur bacteria isolated from an Antarctic microbial mat is described. The organism, designated strain ANT.BR, was mildly psychrophilic, growing optimally at 15-18 degrees C with a growth temperature range of 0-25 degrees C. Cells of strain ANT.BR were highly motile curved rods and spirals, contained bacteriochlorophyll a, and showed a multicomponent in vivo absorption spectrum. A specific phylogenetic relationship was observed between strain ANT.BR and the purple bacterium Rhodoferax fermentans FR2T, and the two organisms shared several physiological and other phenotypic properties, with the notable exception of growth temperature optimum. Tests of genomic DNA hybridization, however, showed Rfx. fermentans FR2T and strain ANT.BR to be genetically distinct bacteria. Because of its unique set of properties, especially its requirement for low growth temperatures, we propose to recognize strain ANT.BR as a new species of the genus Rhodoferax, Rhodoferax antarcticus, named for its known habitat, the Antarctic.

NASA Discipline Exobiology↗

Metabolic versatility of the nitrite-oxidizing bacterium Nitrospira marina and its proteomic response to oxygen-limited conditions

The genus Nitrospira is the most widespread group of nitrite-oxidizing bacteria and thrives in diverse natural and engineered ecosystems. Nitrospira marina Nb-295 T was isolated from the ocean over 30 years ago; however, its genome has not yet been analyzed. Here, we investigated the metabolic potential of N. marina based on its complete genome sequence and performed physiological experiments to test genome-derived hypotheses. Our data confirm that N. marina benefits from additions of undefined organic carbon substrates, has adaptations to resist oxidative, osmotic, and UV light-induced stress and low dissolved p CO 2 , and requires exogenous vitamin B 12 . In addition, N. marina is able to grow chemoorganotrophically on formate, and is thus not an obligate chemolithoautotroph. We further investigated the proteomic response of N. marina to low (~5.6 µM) O 2 concentrations. The abundance of a potentially more efficient CO 2 -fixing pyruvate:ferredoxin oxidoreductase (POR) complex and a high-affinity cbb 3 -type terminal oxidase increased under O 2 limitation, suggesting a role in sustaining nitrite oxidation-driven autotrophy. This putatively more O 2 -sensitive POR complex might be protected from oxidative damage by Cu/Zn-binding superoxide dismutase, which also increased in abundance under low O 2 conditions. Furthermore, the upregulation of proteins involved in alternative energy metabolisms, including Group 3b [NiFe] hydrogenase and formate dehydrogenase, indicate a high metabolic versatility to survive conditions unfavorable for aerobic nitrite oxidation. In summary, the genome and proteome of the first marine Nitrospira isolate identifies adaptations to life in the oxic ocean and provides insights into the metabolic diversity and niche differentiation of NOB in marine environments.

59 BASIC BIOLOGICAL SCIENCES↗

Genome-guided isolation of the hyperthermophilic aerobe Fervidibacter sacchari reveals conserved polysaccharide metabolism in the Armatimonadota

Few aerobic hyperthermophilic microorganisms degrade polysaccharides. Here, we describe the genome-enabled enrichment and optical tweezer-based isolation of an aerobic polysaccharide-degrading hyperthermophile, Fervidibacter sacchari, previously ascribed to candidate phylum Fervidibacteria. F. sacchari uses polysaccharides and monosaccharides for growth at 65–87.5°C and expresses 191 carbohydrate-active enzymes (CAZymes) according to RNA-Seq and proteomics, including 31 with unusual glycoside hydrolase domains (GH109, GH177, GH179). Fluorescence in-situ hybridization and nanoscale secondary ion mass spectrometry confirmed rapid assimilation of 13 C-starch in spring sediments. Purified GHs were optimally active at 80–100°C on ten different polysaccharides. Finally, we propose reassigning Fervidibacteria as a class within phylum Armatimonadota, along with 18 other species, and show that a high number and diversity of CAZymes is a hallmark of the phylum, in both aerobic and anaerobic lineages. Our study establishes Fervidibacteria as hyperthermophilic polysaccharide degraders in terrestrial geothermal springs and suggests a broad role for Armatimonadota in polysaccharide catabolism.

59 BASIC BIOLOGICAL SCIENCES↗

Isolation of genome-predicted Caldatribacterium ( Atribacterota ) reveals pervasive microbial cultivation problem due to folate precipitation

Most bacterial phyla have few or no pure cultures, including Atribacterota , comprised of ubiquitous anaerobes. Here, we report genome-guided enrichment and isolation of two Atribacterota species representing a new family, Caldatribacterium saccharofermentans from a hot spring, and Caldatribacterium inferamans from a deep aquifer. Both were co-enriched with sulfate-reducing bacteria and initially resisted isolation, which we link to inadvertent removal of precipitated folic acid by filter-sterilization of unbuffered Wolin’s vitamin solution. We then predict folate auxotrophy across the Atribacterota and ~29% of all bacteria, with extensive auxotrophy in 27% of phyla. Since ≥604 of 791 ( ≥ 76%) media with folic acid additions in the MediaDive database use unbuffered vitamin solutions in which folic acid is likely removed during filter-sterilization, we propose that folate auxotrophy limits culturability in defined media en masse. We also uncover unusual features of Caldatribacterium , including three lipid membrane-like layers (LMLs), with the inner LML surrounding the nucleoid, and a high percentage of secreted proteins, supporting a unique cell biology of Atribacterota .

Biological and medical sciences↗

Phenylpropanoid methyl esterase unlocks catabolism of aromatic biological nitrification inhibitors

Microbial nitrification of fertilizers represents is a significant global source of greenhouse gas emissions. This process increases emissions, fosters toxic algal blooms, and raises crop production costs. Some plants naturally release biological nitrification inhibitors to suppress ammonium-oxidizing microbes and reduce nitrification. Engineering nitrification inhibitor production into food and bioenergy crops via synthetic biology offers a promising mitigation strategy, but its success depends on addressing gaps in our understanding of inhibitor degradation in soil. This study begins to fill this gap by identifying a previously unknown microbial pathway for degrading phenylpropanoid methyl esters, a key class of aromatic nitrification inhibitors. Using transcriptomics and high-throughput functional genomics, we discovered genes essential for phenylpropanoid methyl ester degradation. Genetic and biochemical analyses revealed two novel enzymes, including a newly identified phenylpropanoid methyl esterase, that direct phenylpropanoid methyl esters into known metabolic pathways. Importantly, transferring these genes into bacteria capable of metabolizing other phenylpropanoids enabled them to use the methyl esters as a carbon source. This work provides critical insights into microbial nitrification inhibitor degradation, a poorly understood element of the nitrification cycle.

Genetic Engineering↗

Listeria monocytogenes requires cellular respiration for NAD+ regeneration and pathogenesis

Cellular respiration is essential for multiple bacterial pathogens and a validated antibiotic target. In addition to driving oxidative phosphorylation, bacterial respiration has a variety of ancillary functions that obscure its contribution to pathogenesis. We find here that the intracellular pathogen Listeria monocytogenes encodes two respiratory pathways which are partially functionally redundant and indispensable for pathogenesis. Loss of respiration decreased NAD + regeneration, but this could be specifically reversed by heterologous expression of a water-forming NADH oxidase (NOX). NOX expression fully rescued intracellular growth defects and increased L. monocytogenes loads >1000-fold in a mouse infection model. Consistent with NAD + regeneration maintaining L. monocytogenes viability and enabling immune evasion, a respiration-deficient strain exhibited elevated bacteriolysis within the host cytosol and NOX expression rescued this phenotype. These studies show that NAD + regeneration represents a major role of L. monocytogenes respiration and highlight the nuanced relationship between bacterial metabolism, physiology, and pathogenesis.

59 BASIC BIOLOGICAL SCIENCES↗

Importance of a martian hematite site for astrobiology

Defining locations where conditions may have been favorable for life is a key objective for the exploration of Mars. Of prime importance are sites where conditions may have been favorable for the preservation of evidence of prebiotic or biotic processes. Areas displaying significant concentrations of the mineral hematite (alpha-Fe2O3), recently identified by thermal emission spectrometry, may have significance in the search for evidence of extraterrestrial life. Since iron oxides can form as aqueous mineral precipitates, the potential exists to preserve microscopic evidence of life in iron oxide-depositing ecosystems. Terrestrial hematite deposits proposed as possible analogs for hematite deposits on Mars include massive (banded) iron formations, iron oxide hydrothermal deposits, iron-rich laterites and ferricrete soils, and rock varnish. We report the potential for long-term preservation of microfossils by iron oxide mineralization in specimens of the approximately 2,100-Ma banded iron deposit of the Gunflint Formation, Canada. Scanning and analytical electron microscopy reveals micrometer-scale rods, spheres, and filaments consisting predominantly of iron and oxygen with minor carbon. We interpret these objects as microbial cells permineralized by an iron oxide, presumably hematite. The confirmation of ancient martian microbial life in hematite deposits will require the return of samples to terrestrial laboratories. A hematite-rich deposit composed of aqueous iron oxide precipitates may thus prove to be a prime site for future sample return.

Exobiology/methods↗

Cannibal's relief: the origins of sex

Microorganisms in the gut of a cockroach resemble those that lived on earth 2000 million years ago. One of their activities, eating each other, may hold the key to the history of sexual reproduction in today's animals and plants.

Non-NASA Center↗

The green non-sulfur bacteria: a deep branching in the eubacterial line of descent

Ribosomal RNA sequence comparisons define a phylogenetic grouping, the green non-sulfur bacteria and relatives (GNS), known to contain the genera Chloroflexus, Herpetosiphon and Thermomicrobium--organisms that have little phenotypic similarity. The unit is phylogenetically deep, but entirely distinct from any other eubacterial division (phylum). It is also relatively ancient--branching from the common eubacterial stem earlier than any other group of eubacteria reported thus far. The group phenotype is predominantly thermophilic, and its thermophilic members, especially Thermomicrobium, are more slowly evolving than Herpetosiphon, a mesophile. The GNS unit appears significantly older than either the green sulfur bacteria or the cyanobacteria--making it likely that organisms such as Chloroflexus, not the cyanobacteria, generated the oldest stromatolites, which formed over three billion years ago.

Non-NASA Center↗

Were the original eubacteria thermophiles?

Thermotoga maritima is one of the more unusual eubacteria: It is highly thermophilic, growing at temperatures higher than any other eubacterium; its cell wall appears to have a unique structure and its lipids a unique composition; and the organism is surrounded by a loose-fitting sheath of unknown function. Its phenotypic uniqueness is matched by its phylogenetic position; Thermotoga maritima represents the deepest known branching in the eubacterial line of descent, as measured by ribosomal RNA sequence comparisons. T. maritima also represents the most slowly evolving of eubacterial lineages. The fact that the two deepest branchings in the eubacterial line of descent (the other, the green non-sulfur bacteria and relatives, i.e. Chloroflexus, Thermomicrobium, etc.) are both basically thermophilic and slowly evolving, strongly suggests that all eubacteria have ultimately arisen from a thermophilic ancestor.

NASA Discipline Exobiology↗