Biomedical subjects

Stephen J Giovannoni

Publications and source records attributed to Stephen J Giovannoni.

At least 19 recordsLinked to original sources

PubMed2006

Pelagibaca bermudensis gen. nov., sp. nov., a novel marine bacterium within the Roseobacter clade in the order Rhodobacterales.

A Gram-negative, chemoheterotrophic, facultatively anaerobic, slightly halophilic, oval-shaped marine bacterium, designated HTCC2601T, was isolated from the western Sargasso Sea by high-throughput culturing involving dilution to extinction. Although the 16S rRNA gene sequence similarity between the isolate and Salipiger mucosus was 96.5 %, phylogenetic analyses using different treeing algorithms clearly indicated that the strain forms a distinct lineage within a clade containing the recently classified genera Salipiger and Palleronia in the order Rhodobacterales of the Alphaproteobacteria. The DNA-DNA relatedness between strain HTCC2601T and S. mucosus was 26.3 %. Strain HTCC2601T utilized a wide range of carbohydrates, including hexose monomers, sugar alcohols, organic acids and amino acids, as sole carbon sources. The DNA G+C content of strain HTCC2601T was 65.4 mol%, and the predominant constituents of the cellular fatty acids were 18 : 1omega7c (79.7 %) and 11-methyl 18 : 1omega7c (7.5 %). The strain differed from members of the closely related genera Salipiger and Palleronia in its morphological, biochemical and ecological characteristics. On the basis of the taxonomic data obtained in this study, a novel genus and species, Pelagibaca bermudensis gen. nov., sp. nov., is proposed; HTCC2601T (=KCTC 12554T = JCM 13377T) is the type strain of Pelagibaca bermudensis.

Molecular Sequence Data↗

PubMed2005

Proteorhodopsin in the ubiquitous marine bacterium SAR11.

Proteorhodopsins are light-dependent proton pumps that are predicted to have an important role in the ecology of the oceans by supplying energy for microbial metabolism. Proteorhodopsin genes were first discovered through the cloning and sequencing of large genomic DNA fragments from seawater. They were later shown to be widely distributed, phylogenetically diverse, and active in the oceans. Proteorhodopsin genes have not been found in cultured bacteria, and on the basis of environmental sequence data, it has not yet been possible to reconstruct the genomes of uncultured bacterial strains that have proteorhodopsin genes. Although the metabolic effect of proteorhodopsins is uncertain, they are thought to function in cells for which the primary mode of metabolism is the heterotrophic assimilation of dissolved organic carbon. Here we report that SAR11 strain HTCC1062 ('Pelagibacter ubique'), the first cultivated member of the extraordinarily abundant SAR11 clade, expresses a proteorhodopsin gene when cultured in autoclaved seawater and in its natural environment, the ocean. The Pelagibacter proteorhodopsin functions as a light-dependent proton pump. The gene is expressed by cells grown in either diurnal light or in darkness, and there is no difference between the growth rates or cell yields of cultures grown in light or darkness.

Alphaproteobacteria↗

PubMed2005

Molecular diversity and ecology of microbial plankton.

The history of microbial evolution in the oceans is probably as old as the history of life itself. In contrast to terrestrial ecosystems, microorganisms are the main form of biomass in the oceans, and form some of the largest populations on the planet. Theory predicts that selection should act more efficiently in large populations. But whether microbial plankton populations harbour organisms that are models of adaptive sophistication remains to be seen. Genome sequence data are piling up, but most of the key microbial plankton clades have no cultivated representatives, and information about their ecological activities is sparse.

Adaptation, Physiological↗

PubMed2005

Genome streamlining in a cosmopolitan oceanic bacterium.

The SAR11 clade consists of very small, heterotrophic marine alpha-proteobacteria that are found throughout the oceans, where they account for about 25% of all microbial cells. Pelagibacter ubique, the first cultured member of this clade, has the smallest genome and encodes the smallest number of predicted open reading frames known for a free-living microorganism. In contrast to parasitic bacteria and archaea with small genomes, P. ubique has complete biosynthetic pathways for all 20 amino acids and all but a few cofactors. P. ubique has no pseudogenes, introns, transposons, extrachromosomal elements, or inteins; few paralogs; and the shortest intergenic spacers yet observed for any cell.

Alphaproteobacteria↗

PubMed2005

Bacterial community composition determined by culture-independent and -dependent methods during propane-stimulated bioremediation in trichloroethene-contaminated groundwater.

An in situ co-metabolic air sparging (CAS) study was carried out at McClellan Air Force Base (MAFB), Sacramento, CA, USA, in a trichloroethene- (TCE) and cis-dichloroethene (cis-DCE)-contaminated aquifer where one test zone received 2% propane in air and the other served as a control and received only air. As part of that study, bacterial population shifts were evaluated by length heterogeneity polymerase chain reaction (LH-PCR). The results showed that an organism(s) that had a fragment size of 385 bp was positively correlated with propane removal rates. The 385 bp fragment consisted of up to 83% of the total fragments in the analysis when propane removal rates peaked. A 16S rRNA clone library made from the bacteria sampled from the propane-sparged groundwater included clones of a TM7 division bacterium that had a 385 bp LH-PCR fragment; no other bacterial species with this fragment size were detected. Both propane removal rates and the 385 bp LH-PCR fragment decreased as nitrate levels in the groundwater decreased. Extinction culturing in natural unamended groundwater medium was used to assess the bacterial diversity of the culturable fraction of microorganisms in both CAS and air-sparged groundwater and to bring novel species into culture for further study. The dominant cultures acquired from the CAS groundwater were from the Herbaspirillum/Oxalobacter clade. The dominant cultures from the air-sparged groundwater were from a novel beta-Proteobacterial clade, which we named after isolate HTCC333.

Bacteria↗

PubMed2004

Oceanicola granulosus gen. nov., sp. nov. and Oceanicola batsensis sp. nov., poly-beta-hydroxybutyrate-producing marine bacteria in the order 'Rhodobacterales'.

Three Gram-negative, chemoheterotrophic, non-motile, rod-shaped bacterial strains that accumulate poly-beta-hydroxybutyrate granules were isolated from the Bermuda Atlantic Time-series Study site by high-throughput culturing methods and characterized by polyphasic approaches. DNA-DNA hybridization, DNA G+C content and phylogenetic analyses based on 16S rRNA gene sequences divided the three isolates into two distinct genospecies that were clearly differentiated by fatty acid profiles, carbon source utilization patterns, antibiotic susceptibility and biochemical characteristics. The strains utilized a wide range of substrates, including pentoses, hexoses, oligosaccharides, sugar alcohols, organic acids and amino acids. DNA G+C contents were 71.5, 70.9 and 67.3 mol% for strains HTCC2516T, HTCC2523 and HTCC2597T, respectively. The most dominant fatty acid was 18 : 1omega7c in strains HTCC2516T and HTCC2523, and cyclo 19 : 0 in strain HTCC2597T. The type strains HTCC2516T and HTCC2597T were clearly differentiated by the presence or absence of 12 : 0, 12 : 1omega11c, 14 : 0, 15 : 0 and methyl 18 : 1. Phylogenetic analyses indicated that the strains formed a distinct monophyletic lineage within the Roseobacter clade in the order 'Rhodobacterales' of the Alphaproteobacteria, and which did not associate with any of the described genera. Genotypic and phenotypic differences of the isolates from the previously described genera support the description of Oceanicola granulosus gen. nov., sp. nov. with the type strain HTCC2516T (=ATCC BAA-861T=DSM 15982T=KCTC 12143T) and of Oceanicola batsensis sp. nov. with the type strain HTCC2597T (=ATCC BAA-863T=DSM 15984T=KCTC 12145T).

Acids↗

PubMed2004

Robiginitalea biformata gen. nov., sp. nov., a novel marine bacterium in the family Flavobacteriaceae with a higher G+C content.

Two Gram-negative, chemoheterotrophic, non-motile, rust-coloured, marine strains were isolated from the western Sargasso Sea by high-throughput culturing. Characterization of the two strains by polyphasic approaches indicated that they are members of the same species. Phylogenetic analyses based on 16S rRNA gene sequences using three treeing algorithms revealed that the strains formed a coherent and novel genus-level lineage within the family Flavobacteriaceae. The dominant fatty acids were branched or hydroxy acids, i15 : 0, i15 : 1 and 3-OH i17 : 0 being the most abundant. The higher DNA G+C content of the strains (55-56 mol%) clearly differentiated them from other genera of the family Flavobacteriaceae (27-44 mol%). It is proposed, from the polyphasic evidence, that the strains be placed into a novel genus and a novel species named Robiginitalea biformata gen. nov., sp. nov., with strain HTCC2501T (=ATCC BAA-864T=KCTC 12146T) as the type strain.

Atlantic Ocean↗

PubMed2004

Lentisphaera araneosa gen. nov., sp. nov, a transparent exopolymer producing marine bacterium, and the description of a novel bacterial phylum, Lentisphaerae.

Two phylogenetically distinct marine strains producing transparent exopolymers (TEP), designated HTCC2155(T) and HTCC2160, were cultivated from Oregon coast seawater by dilution to extinction in a high throughput culturing format. When cultured in low-nutrient seawater media, these strains copiously produced Alcian Blue-stainable viscous TEP. Growing cells were attached to each other by the TEP in a three dimensional network. Polymerase chain reaction employing 16S rDNA primers specific for the novel isolates indicated that they are indigenous to the water column of the Atlantic and Pacific oceans. The abundance of the isolates as determined by 16S rRNA dot blots, however, indicated that they are less than 1% of the total bacterial community. In phylogenetic analyses, the strains consistently formed a new phylum-level lineage within the domain Bacteria, together with members of the candidate phylum VadinBE97, which consists of Victivallis, the first cultured genus in the candidate phylum, and 16S rRNA gene clones from DNA extracted from marine or anaerobic terrestrial habitats. Five putative subgroups were delineated within this phylum-level lineage, including a marine group and an anaerobic group. The isolates are Gram negative, strictly aerobic, chemoheterotrophic, and facultatively oligotrophic sphere-shaped bacteria. The DNA G+C content of strain HTCC2155(T) was 48.3 mol% and the genome size was 2.9 mb. It is proposed from these observations that the strains be placed into a new genus and a new species named Lentisphaera araneosa (type strain HTCC2155(T) = ATCC BAA-859(T) = KCTC 12141(T)) gen. nov., sp. nov., the cultured marine representative of the Lentisphaerae phyl. nov., and the phylum be divided into two novel orders named the Lentisphaerales ord. nov. and the Victivallales ord. nov.

Bacteria↗

PubMed2004

Cultivation and growth characteristics of a diverse group of oligotrophic marine Gammaproteobacteria.

Forty-four novel strains of Gammaproteobacteria were cultivated from coastal and pelagic regions of the Pacific Ocean using high-throughput culturing methods that rely on dilution to extinction in very low nutrient media. Phylogenetic analysis showed that the isolates fell into five rRNA clades, all of which contained rRNA gene sequences reported previously from seawater environmental gene clone libraries (SAR92, OM60, OM182, BD1-7, and KI89A). Bootstrap analyses of phylogenetic reliability did not support collapsing these five clades into a single clade, and they were therefore named the oligotrophic marine Gammaproteobacteria (OMG) group. Twelve cultures chosen to represent the five clades were successively purified in liquid culture, and their growth characteristics were determined at different temperatures and dissolved organic carbon concentrations. The isolates in the OMG group were physiologically diverse heterotrophs, and their physiological properties generally followed their phylogenetic relationships. None of the isolates in the OMG group formed colonies on low- or high-nutrient agar upon their first isolation from seawater, while 7 of 12 isolates that were propagated for laboratory testing eventually produced colonies on 1/10 R2A agar. The isolates grew relatively slowly in natural seawater media (1.23 to 2.63 day(-1)), and none of them grew in high-nutrient media (>351 mg of C liter(-1)). The isolates were psychro- to mesophilic and obligately oligotrophic; many of them were of ultramicrobial size (<0.1 micro m(3)). This cultivation study revealed that sporadically detected Gammaproteobacteria gene clones from seawater are part of a phylogenetically diverse constellation of organisms mainly composed of oligotrophic and ultramicrobial lineages that are culturable under specific cultivation conditions.

Bacteriological Techniques↗

PubMed2004

Representative freshwater bacterioplankton isolated from Crater Lake, Oregon.

High-throughput culturing (HTC) methods that rely on dilution to extinction in very-low-nutrient media were used to obtain bacterial isolates from Crater Lake, Oregon. 16S rRNA sequence determination and phylogenetic reconstruction were used to determine the potential ecological significance of isolated bacteria, both in Crater Lake and globally. Fifty-five Crater Lake isolates yielded 16 different 16S rRNA gene sequences. Thirty of 55 (55%) Crater Lake isolates had 16S rRNA gene sequences with 97% or greater similarity to sequences recovered previously from Crater Lake 16S rRNA gene clone libraries. Furthermore, 36 of 55 (65%) Crater Lake isolates were found to be members of widely distributed freshwater groups. These results confirm that HTC is a significant improvement over traditional isolation techniques that tend to enrich for microorganisms that do not predominate in their environment and rarely correlate with 16S rRNA gene clone library sequences. Although all isolates were obtained under dark, heterotrophic growth conditions, 2 of the 16 different groups showed evidence of photosynthetic capability as assessed by the presence of puf operon sequences, suggesting that photoheterotrophy may be a significant process in this oligotrophic, freshwater habitat.

Actinobacteria↗

PubMed2004

Proteomic analysis of novel marine bacteria using MALDI and ESI mass spectrometry.

The objective of this study was to develop a mass spectrometric protocol to search for proteins related to phototrophy in marine bacteria. The genes that produce proteins involved in conversion of light into energy have been detected by cloning-sequencing from some of these bacteria, but it was previously unknown if these proteins were actually expressed. Attaining this study's goal was complicated by the fact that the samples consisted of miniscule cell pellets, which yielded small amounts of very complex mixtures of proteins. Sample preparation and analysis were tailored to optimize the probability of detecting the proteins of interest. It has been reported that using both matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) to analyze a mixture of peptides leads to the identification of more peptides that either technique alone. In order to exploit this complementarity between ESI and MALDI for proteomic analysis, samples were analyzed using both ionization techniques. With correct choices in sample preparation and ionization process, biologically relevant proteins can be identified out of small samples containing whole proteomes.

Alphaproteobacteria↗

PubMed2003

Fluids from aging ocean crust that support microbial life.

Little is known about the potential for life in the vast, low-temperature (<100 degrees C) reservoir of fluids within mid-ocean ridge flank and ocean basin crust. Recently, an overpressured 300-meter-deep borehole was fitted with an experimental seal (CORK) delivering crustal fluids to the sea floor for discrete and large-volume sampling and characterization. Results demonstrate that the 65 degrees C fluids from 3.5-million-year-old ocean crust support microbial growth. Ribosomal RNA gene sequence data indicate the presence of diverse Bacteria and Archaea, including gene clones of varying degrees of relatedness to known nitrate reducers (with ammonia production), thermophilic sulfate reducers, and thermophilic fermentative heterotrophs, all consistent with fluid chemistry.

Ammonia↗

PubMed2003

Croceibacter atlanticus gen. nov., sp. nov., a novel marine bacterium in the family Flavobacteriaceae.

A bright, saffron-colored marine bacterium HTCC2559T was isolated from the Bermuda Atlantic Time Series station in the western Sargasso Sea, Atlantic Ocean by high throughput culturing methods and characterized by polyphasic approaches. Phenotypic data and phylogenetic analyses showed that the strain is a member of the family Flavobacteriaceae. The strain was gram-negative, non-motile, chemoheterotrophic, strictly aerobic, NaCl-requiring, rod-shaped cells that contain carotenoid pigments but not flexirubin. Several kinds of macromolecules (gelatin, DNA, starch, casein, and elastin) were degraded and carbohydrates, sugar alcohols, organic acids, and amino acids were utilized as sole carbon sources. The dominant fatty acids were branched or hydroxy acids, and 3-OH i17:0, i15:0, i15:1, and i17:1 omega9c were abundant. The DNA G+C content of the strain is 34.8 mol%. Phylogenetic analyses using three treeing algorithms based on 16S rRNA gene sequences revealed that the strain formed a very distinct lineage that is allied closely with several seawater environmental clones in the family Flavobacteriaceae. Therefore, it is proposed from the polyphasic studies that strain HTCC2559T (=ATCC BAA-628T = KCTC 12090T) belongs to a new genus and species named Croceibacter atlanticus gen. nov., sp. nov.

Biodegradation, Environmental↗

PubMed2003

Parvularcula bermudensis gen. nov., sp. nov., a marine bacterium that forms a deep branch in the alpha-Proteobacteria.

Two bacterial strains, HTCC2503T and HTCC2517, were isolated from the Bermuda Atlantic Time Series Station in the western Sargasso Sea, Atlantic Ocean, by new high-throughput culture methods that rely on dilution to extinction in very-low-nutrient media. Characterization of the two strains by polyphasic approaches revealed that they belonged to the same species. These isolates are gram-negative, strictly aerobic, chemoheterotrophic, slightly motile short rods with a single flagellum. The temperature, pH and NaCl concentration ranges for growth were 10-37 degrees C, 6.0-9.0 and 0.75-20% (w/v), respectively. Colonies on marine agar were very small (0.3-0.8 mm in diameter), yellowish-brown and very hard. Carotenoid pigments were synthesized but bacteriochlorophyll a was not. Several kinds of pentose, hexose, sugar alcohol, oligosaccharide and amino acid were utilized as sole carbon sources. Oxidase was produced, but catalase was not. All cellular fatty acids were even-numbered monounsaturated or saturated fatty acids and the major fatty acid was cis-7-octadecenoic acid (73.3%). The DNA G + C content of strain HTCC2503T was 60.8 mol%. Phylogenetic analyses of 16S rRNA gene sequences clearly indicated that the strains formed a distinct lineage, allied with activated sludge environmental clone H9, in the alpha-Proteobacteria. The clade containing strains HTCC2503T and HTCC2517 and clone H9 could not be phylogenetically associated with any of the six known orders of the alpha-Proteobacteria. From this polyphasic evidence, it is proposed that the novel strains should be classified as Parvularcula bermudensis gen. nov., sp. nov. The type strain is HTCC2503T (= ATCC BAA-594T = KCTC 12087T) and the reference strain is HTCC2517.

Alphaproteobacteria↗

PubMed2003

Fulvimarina pelagi gen. nov., sp. nov., a marine bacterium that forms a deep evolutionary lineage of descent in the order "Rhizobiales".

Three brownish-yellow bacterial strains were isolated from the western Sargasso Sea by high-throughput culturing methods and characterized by polyphasic approaches. All isolates were Gram-negative, strictly aerobic, chemoheterotrophic, non-motile short rods that contained carotenoid pigments. Phylogenetic analyses based on 16S rRNA gene sequences, DNA-DNA hybridization and DNA G+C content, along with phenotypic characteristics, revealed that they belonged to the same species. The strains utilized a wide range of substrates, including pentoses, hexoses, oligosaccharides, sugar alcohols, organic acids and amino acids, as sole carbon sources. The DNA G+C content of the isolates ranged from 57.6 to 59.9 mol%. The predominant cellular fatty acid constituent was C(18 : 1)omega7c, whilst C(16 : 0), C(18 : 0) and C(19 : 0)omega8c cyclo were also abundant. The organism related most closely to these strains, as determined by 16S rDNA sequence comparison, was the recently described species Aurantimonas coralicida (93.3-93.8 % similarity). Phylogenetic analyses indicated that the strains formed a distinct and deep evolutionary lineage of descent, together with A. coralicida, within the order "Rhizobiales" of the alpha-Proteobacteria. This lineage could not be associated with any of the ten known families in the order "Rhizobiales". From polyphasic evidence, it is proposed that the strains be placed into a novel genus and species, Fulvimarina pelagi gen. nov., sp. nov. (type strain, HTCC2506(T)=ATCC BAA-666(T)=KCTC 12091(T)=DSM 15513(T)).

Microbial Sensitivity Tests↗

PubMed2003

The uncultured microbial majority.

Since the delineation of 12 bacterial phyla by comparative phylogenetic analyses of 16S ribosomal RNA in 1987 knowledge of microbial diversity has expanded dramatically owing to the sequencing of ribosomal RNA genes cloned from environmental DNA. Currently, only 26 of the approximately 52 identifiable major lineages, or phyla, within the domain Bacteria have cultivated representatives. Evidence from field studies indicates that many of the uncultivated phyla are found in diverse habitats, and some are extraordinarily abundant. In some important environments, including seawater, freshwater, and soil, many biologically and geochemically important organisms are at best only remotely related to any strain that has been characterized by phenotype or by genome sequencing. Genome sequence information that would allow ribosomal RNA gene trees to be related to broader patterns in microbial genome evolution is scant, and therefore microbial diversity remains largely unexplored territory.

DNA, Bacterial↗

PubMed2002

Cultivation of the ubiquitous SAR11 marine bacterioplankton clade.

The alpha-proteobacterial lineage that contains SAR11 and related ribosomal RNA gene clones was among the first groups of organisms to be identified when cultivation-independent approaches based on rRNA gene cloning and sequencing were applied to survey microbial diversity in natural ecosystems. This group accounts for 26% of all ribosomal RNA genes that have been identified in sea water and has been found in nearly every pelagic marine bacterioplankton community studied by these methods. The SAR11 clade represents a pervasive problem in microbiology: despite its ubiquity, it has defied cultivation efforts. Genetic evidence suggests that diverse uncultivated microbial taxa dominate most natural ecosystems, which has prompted widespread efforts to elucidate the geochemical activities of these organisms without the benefit of cultures for study. Here we report the isolation of representatives of the SAR11 clade. Eighteen cultures were initially obtained by means of high-throughput procedures for isolating cell cultures through the dilution of natural microbial communities into very low nutrient media. Eleven of these cultures have been successfully passaged and cryopreserved for future study. The volume of these cells, about 0.01 micro m(3), places them among the smallest free-living cells in culture.

Alphaproteobacteria↗

PubMed2002

High-throughput methods for culturing microorganisms in very-low-nutrient media yield diverse new marine isolates.

Microbial diversity studies based on the cloning and sequencing of DNA from nature support the conclusion that only a fraction of the microbial diversity is currently represented in culture collections. Out of over 40 known prokaryotic phyla, only half have cultured representatives. In an effort to culture the uncultured phylotypes from oligotrophic marine ecosystems, we developed high-throughput culturing procedures that utilize the concept of extinction culturing to isolate cultures in small volumes of low-nutrient media. In these experiments, marine bacteria were isolated and cultivated at in situ substrate concentrations-typically 3 orders of magnitude less than common laboratory media. Microtiter plates and a newly developed procedure for making cell arrays were employed to raise the throughput rate and lower detection sensitivity, permitting cell enumeration from 200-microl aliquots of cultures with densities as low as 10(3) cells/ml. Approximately 2,500 extinction cultures from 11 separate samplings of marine bacterioplankton were screened over the course of 3 years. Up to 14% of the cells collected from coastal seawater were cultured by this method, which was 14- to 1,400-fold higher than the numbers obtained by traditional microbiological culturing techniques. Among the microorganisms cultured were four unique cell lineages that belong to previously uncultured or undescribed marine Proteobacteria clades known from environmental gene cloning studies. These cultures are related to the clades SAR11 (alpha subclass), OM43 (beta subclass), SAR92 (gamma subclass), and OM60/OM241 (gamma subclass). This method proved successful for the cultivation of previously uncultured marine bacterioplankton that have consistently been found in marine clone libraries.

Bacteriological Techniques↗