bluuebunny/crossref_metadata_2025_split huggingface dataset:
Stable-Isotope Probing of Microorganisms Thriving at Thermodynamic Limits: Syntrophic Propionate Oxidation in Flooded Soil
Authors: Lueders Tillmann , Pommerenke Bianca , Friedrich Michael W. 
Published: October 2004

ABSTRACT

Propionate is an important intermediate of the degradation of organic matter in many anoxic environments. In methanogenic environments, due to thermodynamic constraints, the oxidation of propionate requires syntrophic cooperation of propionate-fermenting proton-reducing bacteria and H
2
-consuming methanogens. We have identified here microorganisms that were active in syntrophic propionate oxidation in anoxic paddy soil by rRNA-based stable-isotope probing (SIP). After 7 weeks of incubation with [
13
C]propionate (<10 mM) and the oxidation of ∼30 μmol of
13
C-labeled substrate per g dry weight of soil, we found that archaeal nucleic acids were
13
C labeled to a larger extent than those of the bacterial partners. Nevertheless, both terminal restriction fragment length polymorphism and cloning analyses revealed
Syntrophobacter
spp.,
Smithella
spp., and the novel
Pelotomaculum
spp. to predominate in “heavy”
13
C-labeled bacterial rRNA, clearly showing that these were active in situ in syntrophic propionate oxidation. Among the
Archaea
, mostly
Methanobacterium
and
Methanosarcina
spp. and also members of the yet-uncultured “rice cluster I” lineage had incorporated substantial amounts of
13
C label, suggesting that these methanogens were directly involved in syntrophic associations and/or thriving on the [
13
C]acetate released by the syntrophs. With this first application of SIP in an anoxic soil environment, we were able to clearly demonstrate that even guilds of microorganisms growing under thermodynamic constraints, as well as phylogenetically diverse syntrophic associations, can be identified by using SIP. This approach holds great promise for determining the structure and function relationships of further syntrophic or other nutritional associations in natural environments and for defining metabolic functions of yet-uncultivated microorganisms.
Lactate-Utilizing Bacteria, Isolated from Human Feces, That Produce Butyrate as a Major Fermentation Product
Authors: Duncan Sylvia H. , Louis Petra , Flint Harry J. 
Published: October 2004

ABSTRACT

The microbial community of the human colon contains many bacteria that produce lactic acid, but lactate is normally detected only at low concentrations (<5 mM) in feces from healthy individuals. It is not clear, however, which bacteria are mainly responsible for lactate utilization in the human colon. Here, bacteria able to utilize lactate and produce butyrate were identified among isolates obtained from 10
−8
dilutions of fecal samples from five different subjects. Out of nine such strains identified, four were found to be related to
Eubacterium hallii
and two to
Anaerostipes caccae
, while the remaining three represent a new species within clostridial cluster XIVa based on their 16S rRNA sequences. Significant ability to utilize lactate was not detected in the butyrate-producing species
Roseburia intestinalis
,
Eubacterium rectale
, or
Faecalibacterium prausnitzii
. Whereas
E. hallii
and
A. caccae
strains used both
d
- and
l
-lactate, the remaining strains used only the
d
form. Addition of glucose to batch cultures prevented lactate utilization until the glucose became exhausted. However, when two
E. hallii
strains and one
A. caccae
strain were grown in separate cocultures with a starch-utilizing
Bifidobacterium adolescentis
isolate, with starch as the carbohydrate energy source, the
l
-lactate produced by
B. adolescentis
became undetectable and butyrate was formed. Such cross-feeding may help to explain the reported butyrogenic effect of certain dietary substrates, including resistant starch. The abundance of
E. hallii
in particular in the colonic ecosystem suggests that these bacteria play important roles in preventing lactate accumulation.
Identification of the Receptor-Binding Protein in 936-Species Lactococcal Bacteriophages
Authors: Dupont Kitt , Vogensen Finn Kvist , Neve Horst , Bresciani José , Josephsen Jytte 
Published: October 2004

ABSTRACT

The aim of this work was to identify genes responsible for host recognition in the lactococcal phages sk1 and bIL170 belonging to species 936. These phages have a high level of DNA identity but different host ranges. Bioinformatic analysis indicated that homologous genes,
orf18
in sk1 and
orf20
in bIL170, could be the receptor-binding protein (RBP) genes, since the resulting proteins were unrelated in the C-terminal part and showed homology to different groups of proteins hypothetically involved in host recognition. Consequently, chimeric bIL170 phages carrying
orf18
from sk1 were generated. The recombinant phages were able to form plaques on the sk1 host
Lactococcus lactis
MG1614, and recombination was verified by PCR analysis directly with the plaques. A polyclonal antiserum raised against the C-terminal part of phage sk1 ORF18 was used in immunogold electron microscopy to demonstrate that ORF18 is located at the tip of the tail. Sequence analysis of corresponding proteins from other lactococcal phages belonging to species 936 showed that the N-terminal parts of the RBPs were very similar, while the C-terminal parts varied, suggesting that the C-terminal part plays a role in receptor binding. The phages investigated could be grouped into sk1-like phages (p2, fd13, jj50, and φ7) and bIL170-like phages (P008, P113G, P272, and bIL66) on the basis of the homology of their RBPs to the C-terminal part of ORF18 in sk1 and ORF20 in bIL170, respectively. Interestingly, sk1-like phages bind to and infect a defined group of
L. lactis
subsp.
cremoris
strains, while bIL170-like phages bind to and infect a defined group of
L. lactis
subsp.
lactis
strains.
Identification of
Lactococcus lactis
Genes Required for Bacteriophage Adsorption
Authors: Dupont Kitt , Janzen Thomas , Vogensen Finn Kvist , Josephsen Jytte , Stuer-Lauridsen Birgitte 
Published: October 2004

ABSTRACT

The aim of this work was to identify genes in
Lactococcus lactis
subsp.
lactis
IL1403 and
Lactococcus lactis
subsp.
cremoris
Wg2 important for adsorption of the 936-species phages bIL170 and φ645, respectively. Random insertional mutagenesis of the two
L. lactis
strains was carried out with the vector pGh9:IS
S1
, and integrants that were resistant to phage infection and showed reduced phage adsorption were selected. In
L. lactis
IL1403 integration was obtained in the
ycaG
and
rgpE
genes, whereas in
L. lactis
Wg2 integration was obtained in two genes homologous to
ycbC
and
ycbB
of
L. lactis
IL1403.
rgpE
and
ycbB
encode putative glycosyltransferases, whereas
ycaG
and
ycbC
encode putative membrane-spanning proteins with unknown functions. Interestingly,
ycaG
,
rgpE
,
ycbC
, and
ycbB
are all part of the same operon in
L. lactis
IL1403. This operon is probably involved in biosynthesis and transport of cell wall polysaccharides (WPS). Binding and infection studies showed that φ645 binds to and infects
L. lactis
Wg2,
L. lactis
IL1403, and
L. lactis
IL1403 strains with pGh9:IS
S1
integration in
ycaG
and
rgpE
, whereas bIL170 binds to and infects only
L. lactis
IL1403 and cannot infect Wg2. These results indicate that φ645 binds to a WPS structure present in both
L. lactis
IL1403 and
L. lactis
Wg2, whereas bIL170 binds to another WPS structure not present in
L. lactis
Wg2. Binding of bIL170 and φ645 to different WPS structures was supported by alignment of the receptor-binding proteins of bIL170 and φ645 that showed no homology in the C-terminal part.
Functional Analysis of Fructosyl-Amino Acid Oxidases of
Aspergillus oryzae
Authors: Akazawa Shin-ichi , Karino Tetsuya , Yoshida Nobuyuki , Katsuragi Tohoru , Tani Yoshiki 
Published: October 2004

ABSTRACT

Three active fractions of fructosyl-amino acid oxidase (FAOD-Ao1, -Ao2a, and -Ao2b) were isolated from
Aspergillus oryzae
strain RIB40. N-terminal and internal amino acid sequences of FAOD-Ao2a corresponded to those of FAOD-Ao2b, suggesting that these two isozymes were derived from the same protein. FAOD-Ao1 and -Ao2 were different in substrate specificity and subunit assembly; FAOD-Ao2 was active toward
N
ε
-fructosyl
N
α
-Z-lysine and fructosyl valine (Fru-Val), whereas FAOD-Ao1 was not active toward Fru-Val. The genes encoding the FAOD isozymes (i.e.,
FAOAo1
and
FAOAo2
) were cloned by PCR with an FAOD-specific primer set. The deduced amino acid sequences revealed that FAOD-Ao1 was 50% identical to FAOD-Ao2, and each isozyme had a peroxisome-targeting signal-1, indicating their localization in peroxisomes. The genes was expressed in
Escherichia coli
and rFaoAo2 showed the same characteristics as FAOD-Ao2, whereas rFaoAo1 was not active.
FAOAo2
disruptant was obtained by using
ptrA
as a selective marker. Wild-type strain grew on the medium containing Fru-Val as the sole carbon and nitrogen sources, but strain
ΔfaoAo2
did not grow. Addition of glucose or (NH
4
)
2
SO
4
to the Fru-Val medium did not affect the assimilation of Fru-Val by wild-type, indicating glucose and ammonium repressions did not occur in the expression of the
FAOAo2
gene. Furthermore, conidia of the wild-type strain did not germinate on the medium containing Fru-Val and NaNO
2
as the sole carbon and nitrogen sources, respectively, suggesting that Fru-Val may also repress gene expression of nitrite reductase. These results indicated that FAOD is needed for utilization of fructosyl-amino acids as nitrogen sources in
A. oryzae
.
Oxygen- and Glucose-Dependent Regulation of Central Carbon Metabolism inPichia anomala
Authors: Fredlund Elisabeth , Blank Lars M. , Schnürer Johan , Sauer Uwe , Passoth Volkmar 
Published: October 2004

ABSTRACTWe investigated the regulation of the central aerobic and hypoxic metabolism of the biocontrol and non-Saccharomyceswine yeastPichia anomala. In aerobic batch culture,P. anomalagrows in the respiratory mode with a high biomass yield (0.59 g [dry weight] of cells g of glucose−1) and marginal ethanol, glycerol, acetate, and ethyl acetate production. Oxygen limitation, but not glucose pulse, induced fermentation with substantial ethanol production and 10-fold-increased ethyl acetate production. Despite low or absent ethanol formation, the activities of pyruvate decarboxylase and alcohol dehydrogenase were high during aerobic growth on glucose or succinate. No activation of these enzyme activities was observed after a glucose pulse. However, after the shift to oxygen limitation, both enzymes were activated threefold. Metabolic flux analysis revealed that the tricarboxylic acid pathway operates as a cycle during aerobic batch culture and as a two-branched pathway under oxygen limitation. Glucose catabolism through the pentose phosphate pathway was lower during oxygen limitation than under aerobic growth. Overall, our results demonstrate thatP. anomalaexhibits a Pasteur effect and not a Crabtree effect, i.e., oxygen availability, but not glucose concentration, is the main stimulus for the regulation of the central carbon metabolism.
Evaluation of Methods for Storage of Marine Macroorganisms with Optimal Recovery of Bacteria
Authors: Siebert Kathrin , Busl Martina , Asmus Irina , Freund Josef , Muscholl-Silberhorn Albrecht , Wirth Reinhard 
Published: October 2004

ABSTRACT
Marine macroorganisms are a potential source for new bioactive substances. In many cases marine microorganisms—especially bacteria—associated with these macroorganisms are actually producing the bioactive substances. One often is not able to immediately isolate microorganisms from collected macroorganismic materials; we therefore evaluated different methods for storage of such material, e.g., on board research vessels. These methods were the following: storage of macerates in sintered glass beads and 5% trehalose at −20°C (SGT method); storage of sections in 5% dimethyl sulfoxide at −70°C (SD method); storage of macerates at −20°C using the commercial ROTI-STORE system (RS method); storage of macerates at −20°C in 50% glycerol (GC method); and storage of macerates covered by mineral oil at 4°C (MO method). The SGT and SD methods resulted in numbers of and especially diversity of recoverable bacteria that were higher than for the other methods. Data for the RS method indicated its potential usefulness, too. The MO method resulted in growth during storage, thereby enriching a few selected microorganisms; the GC method resulted in a survival and diversity of recovered bacteria that was too low.
Functional Expression of
Sinorhizobium meliloti
BetS, a High-Affinity Betaine Transporter, in
Bradyrhizobium japonicum
USDA110
Authors: Boscari Alexandre , Mandon Karine , Poggi Marie-Christine , Le Rudulier Daniel 
Published: October 2004

ABSTRACT

Among the
Rhizobiaceae
,
Bradyrhizobium japonicum
strain USDA110 appears to be extremely salt sensitive, and the presence of glycine betaine cannot restore its growth in medium with an increased osmolarity (E. Boncompagni, M. Østerås, M. C. Poggi, and D. Le Rudulier, Appl. Environ. Microbiol. 65:2072-2077, 1999). In order to improve the salt tolerance of
B. japonicum
, cells were transformed with the
betS
gene of
Sinorhizobium meliloti
. This gene encodes a major glycine betaine/proline betaine transporter from the betaine choline carnitine transporter family and is required for early osmotic adjustment. Whereas betaine transport was absent in the USDA110 strain, such transformation induced glycine betaine and proline betaine uptake in an osmotically dependent manner. Salt-treated transformed cells accumulated large amounts of glycine betaine, which was not catabolized. However, the accumulation was reversed through rapid efflux during osmotic downshock. An increased tolerance of transformant cells to a moderate NaCl concentration (80 mM) was also observed in the presence of glycine betaine or proline betaine, whereas the growth of the wild-type strain was totally abolished at 80 mM NaCl. Surprisingly, the deleterious effect due to a higher salt concentration (100 mM) could not be overcome by glycine betaine, despite a significant accumulation of this compound. Cell viability was not significantly affected in the presence of 100 mM NaCl, whereas 75% cell death occurred at 150 mM NaCl. The absence of a potential gene encoding Na
+
/H
+
antiporters in
B. japonicum
could explain its very high Na
+
sensitivity.
Vulnerability of Drinking-Water Wells in La Crosse, Wisconsin, to Enteric-Virus Contamination from Surface Water Contributions
Authors: Borchardt Mark A. , Haas Nathaniel L. , Hunt Randall J. 
Published: October 2004

ABSTRACT

Human enteric viruses can contaminate municipal drinking-water wells, but few studies have examined the routes by which viruses enter these wells. In the present study, the objective was to monitor the municipal wells of La Crosse, Wisconsin, for enteric viruses and determine whether the amount of Mississippi River water infiltrating the wells was related to the frequency of virus detection. From March 2001 to February 2002, one river water site and four wells predicted by hydrogeological modeling to have variable degrees of surface water contributions were sampled monthly for enteric viruses, microbial indicators of sanitary quality, and oxygen and hydrogen isotopes.
18
O/
16
O and
2
H/
1
H ratios were used to determine the level of surface water contributions. All samples were collected prior to chlorination at the wellhead. By reverse transcription-PCR (RT-PCR), 24 of 48 municipal well water samples (50%) were positive for enteric viruses, including enteroviruses, rotavirus, hepatitis A virus (HAV), and noroviruses. Of 12 river water samples, 10 (83%) were virus positive by RT-PCR. Viable enteroviruses were not detected by cell culture in the well samples, although three well samples were positive for culturable HAV. Enteroviruses detected in the wells by RT-PCR were identified as several serotypes of echoviruses and group A and group B coxsackieviruses. None of the well water samples was positive for indicators of sanitary quality, namely male-specific and somatic coliphages, total coliform bacteria,
Escherichia coli
, and fecal enterococci. Contrary to expectations, viruses were found in all wells regardless of the level of surface water contributions. This result suggests that there were other unidentified sources, in addition to surface water, responsible for the contamination.
High-Level Genotypic Variation and Antibiotic Sensitivity among
Escherichia coli
O157 Strains Isolated from Two Scottish Beef Cattle Farms
Authors: Vali Leila , Wisely Karen A. , Pearce Michael C. , Turner Esther J. , Knight Hazel I. , Smith Alastair W. , Amyes Sebastian G. B. 
Published: October 2004

ABSTRACT

Escherichia coli
O157:H7 is a human pathogen that is carried and transmitted by cattle. Scotland is known to have one of the highest rates of
E. coli
O157 human infections in the world. Two hundred ninety-three isolates were obtained from naturally infected cattle and the environment on two farms in the Scottish Highlands. The isolates were typed by pulsed-field gel electrophoresis (PFGE) with XbaI restriction endonuclease enzyme, and 19 different variations in patterns were found. There was considerable genomic diversity within the
E. coli
O157 population on the two farms. The PFGE pattern of one of the observed subtypes matched exactly with that of a strain obtained from a Scottish patient with hemolytic-uremic syndrome. To examine the stability of an individual
E. coli
O157 strain, continuous subculturing of a strain was performed 110 times. No variation from the original PFGE pattern was observed. We found three indistinguishable subtypes of
E. coli
O157 on both study farms, suggesting common sources of infection. We also examined the antibiotic resistance of the isolated strains. Phenotypic studies demonstrated resistance of the strains to sulfamethoxazole (100%), chloramphenicol (3.07%), and at a lower rate, other antibiotics, indicating the preservation of antibiotic sensitivity in a rapidly changing population of
E. coli
O157.
Molecular Analysis of Geographic Patterns of Eukaryotic Diversity in Antarctic Soils
Authors: Lawley Blair , Ripley Sarah , Bridge Paul , Convey Peter 
Published: October 2004

ABSTRACT
We describe the application of molecular biological techniques to estimate eukaryotic diversity (primarily fungi, algae, and protists) in Antarctic soils across a latitudinal and environmental gradient between approximately 60 and 87°S. The data were used to (i) test the hypothesis that diversity would decrease with increasing southerly latitude and environmental severity, as is generally claimed for “higher” faunal and plant groups, and (ii) investigate the level of endemicity displayed in different taxonomic groups. Only limited support was obtained for a systematic decrease in diversity with latitude, and then only at the level of a gross comparison between maritime (Antarctic Peninsula/Scotia Arc) and continental Antarctic sites. While the most southerly continental Antarctic site was three to four times less diverse than all maritime sites, there was no evidence for a trend of decreasing diversity across the entire range of the maritime Antarctic (60 to 72°S). Rather, we found the reverse pattern, with highest diversity at sites on Alexander Island (ca. 72°S), at the southern limit of the maritime Antarctic. The very limited overlap found between the eukaryotic biota of the different study sites, combined with their generally low relatedness to existing sequence databases, indicates a high level of Antarctic site isolation and possibly endemicity, a pattern not consistent with similar studies on other continents.
Enzymatic Analysis of an Amylolytic Enzyme from the Hyperthermophilic Archaeon
Pyrococcus furiosus
Reveals Its Novel Catalytic Properties as both an α-Amylase and a Cyclodextrin-Hydrolyzing Enzyme
Authors: Yang Sung-Jae , Lee Hee-Seob , Park Cheon-Seok , Kim Yong-Ro , Moon Tae-Wha , Park Kwan-Hwa 
Published: October 2004

ABSTRACT

Genomic analysis of the hyperthermophilic archaeon
Pyrococcus furiosus
revealed the presence of an open reading frame (ORF PF1939) similar to the enzymes in glycoside hydrolase family 13. This amylolytic enzyme, designated PFTA (
Pyrococcus furiosus
thermostable amylase), was cloned and expressed in
Escherichia coli
. The recombinant PFTA was extremely thermostable, with an optimum temperature of 90°C. The substrate specificity of PFTA suggests that it possesses characteristics of both α-amylase and cyclodextrin-hydrolyzing enzyme. Like typical α-amylases, PFTA hydrolyzed maltooligosaccharides and starch to produce mainly maltotriose and maltotetraose. However, it could also attack and degrade pullulan and β-cyclodextrin, which are resistant to α-amylase, to primarily produce panose and maltoheptaose, respectively. Furthermore, acarbose, a potent α-amylase inhibitor, was drastically degraded by PFTA, as is typical of cyclodextrin-hydrolyzing enzymes. These results confirm that PFTA possesses novel catalytic properties characteristic of both α-amylase and cyclodextrin-hydrolyzing enzyme.
Potential Role of a Novel Psychrotolerant Member of the FamilyGeobacteraceae,Geopsychrobacter electrodiphilusgen. nov., sp. nov., in Electricity Production by a Marine Sediment Fuel Cell
Authors: Holmes Dawn E. , Nicoll Julie S. , Bond Daniel R. , Lovley Derek R. 
Published: October 2004

ABSTRACTPrevious studies have shown that members of the familyGeobacteraceaethat attach to the anodes of sediment fuel cells are directly involved in harvesting electricity by oxidizing organic compounds to carbon dioxide and transferring the electrons to the anode. In order to learn more about this process, microorganisms from the anode surface of a marine sediment fuel cell were enriched and isolated with Fe(III) oxide. Two unique marine isolates were recovered, strains A1Tand A2. They are gram-negative, nonmotile rods, with abundantc-type cytochromes. Phylogenetic analysis of the 16S rRNA,recA,gyrB,fusA,rpoB, andnifDgenes indicated that strains A1Tand A2 represent a unique phylogenetic cluster within theGeobacteraceae. Both strains were able to grow with an electrode serving as the sole electron acceptor and transferred ca. 90% of the electrons available in their organic electron donors to the electrodes. These organisms are the first psychrotolerant members of theGeobacteraceaereported thus far and can grow at temperatures between 4 and 30°C, with an optimum temperature of 22°C. Strains A1Tand A2 can utilize a wide range of traditional electron acceptors, including all forms of soluble and insoluble Fe(III) tested, anthraquinone 2,6-disulfonate, and S0. In addition to acetate, both strains can utilize a number of other organic acids, amino acids, long-chain fatty acids, and aromatic compounds to support growth with Fe(III) nitrilotriacetic acid as an electron acceptor. The metabolism of these organisms differs in that only strain A1Tcan use acetoin, ethanol, and hydrogen as electron donors, whereas only strain A2 can use lactate, propionate, and butyrate. The nameGeopsychrobacter electrodiphilusgen. nov., sp. nov., is proposed for strains A1Tand A2, with strain A1T(ATCC BAA-880T; DSM 16401T; JCM 12469) as the type strain. Strains A1Tand A2 (ATCC BAA-770; JCM 12470) represent the first organisms recovered from anodes that can effectively couple the oxidation of organic compounds to an electrode. Thus, they may serve as important model organisms for further elucidation of the mechanisms of microbe-electrode electron transfer in sediment fuel cells.