bluuebunny/crossref_metadata_2025 huggingface dataset:
4,5-Propylenedithio-1,3-dithiole-2-thione and 4,5-propylenedithio-1,3-dithiol-2-one
Authors: Xue Gang , Fang Qi , Yu Wentao
Published: November 2003

4,5-Propylenedithio-1,3-dithiole-2-thione, C6H6S5, (I), crystallizes in the centrosymmetric space group P21/c. The molecular packing is characterized by pairs of S...S intermolecular contacts between neighbouring molecules, which may account for the rather high thermal stablity of the crystal. 4,5-Propylenedithio-1,3-dithiol-2-one, C6H6OS4, (II), in which an O atom replaces the terminal S atom of (I), crystallizes in the non-centrosymmetric polar space group Cc. The packing pattern of (II) indicates that the macropolarization direction is along [101]. Although the packing patterns are qualitatively significantly different, the molecular structures of (I) and (II) are similar, each exhibiting a chair conformation.
Pseudosymmetry in ammonium [(carboxymethyl)sulfanyl]acetate
Authors: Maurin Jan K. , Pawłowski Michał , Czarnocki Zbigniew
Published: November 2003

The structure of the title compound, NH4
+·C4H5O4S−, is composed of monocarboxylate anions of [(carboxymethyl)sulfanyl]acetic acid linked into infinite chains via strong O—H...O− hydrogen bonds. The three-dimensional structure is completed by the ammonium cations, which interlink neighbouring chains via N—H...O hydrogen bonds. Solution and refinement in the true space group Pn led to an unambiguous position for the single carboxyl H atom. In the higher symmetry space group P2/n, the carboxylate anion would be located on a twofold axis.
Development of executive attention in preschool children
Authors: Jones Laura Backen , Rothbart Mary K. , Posner Michael I.
Published: November 2003

The development of self‐regulatory systems during the preschool years is accompanied by a dramatic increase in the ability to inhibit actions based on the directions of others. Several tasks have shown evidence of changes in self‐regulation during the fourth year of life. The current cross‐sectional study used a Simple Simon task, in which 33 3‐ to 4‐year‐old children were asked to respond to the command of one large toy animal but not to the command of another. Three important aspects of self‐regulation were examined: the ability to inhibit action in the face of conflict, error detection/correction and the use of verbal and physical control strategies. The ability to inhibit a response in this task increased from 22% to 90% between 36 and 48 months of age. Post error slowing of Reaction Time (RT) indicative of error detection emerged at about the same age as successful inhibition. Physical rather than verbal self‐regulation strategies were spontaneously employed by children to aid in the process of inhibition.
Multiple causality in developmental disorders: methodological implications from computational modelling
Authors: Thomas Michael S.C.
Published: November 2003

When developmental disorders are defined on the basis of behavioural impairments alone, there is a risk that individuals with different underlying cognitive deficits will be grouped together on the basis that they happen to share a certain impairment. This phenomenon is labelledmultiple causality. In contrast, a developmental disorder generated by a single underlying cognitive deficit may nevertheless show variable patterns of impairments due to individual differences. Connectionist computational models of development are used to investigate whether there may be ways to distinguish disorder groups with a single underlying cause (homogeneous disorder groups) from disorder groups with multiple underlying causes (heterogeneous disorder groups) on the basis of behavioural measures alone. A heuristic is proposed to assess the underlying causal homogeneity of the disorder group based on the variability of different behavioural measures from the target domain. Heterogeneous disorder groups are likely to show smaller variability on the measure used to define the disorder than on subsequent behavioural measures, while homogeneous groups should show approximately equivalent variability. Homogeneous disorder groups should show reductions in the variability of behavioural measures over time, while heterogeneous groups may not. It is demonstrated how these predictions arise from computational assumptions, and their use is illustrated with reference to behavioural data on naming skills from two developmental disorder groups, Williams syndrome and children with Word Finding Difficulties.
Testing the abstractness of children's linguistic representations: lexical and structural priming of syntactic constructions in young children
Authors: Savage Ceri , Lieven Elena , Theakston Anna , Tomasello Michael
Published: November 2003

The current studies used a priming methodology to assess the abstractness of children's early syntactic constructions. In the main study, 3‐, 4‐ and 6‐year‐old children were asked to describe a prime picture by repeating either an active or a passive sentence, and then they were left to their own devices to describe a target picture. For half the children at each age, the prime sentences they repeated had high lexical overlap with the sentence they were likely to produce for the target, whereas for the other half there was very low lexical overlap between prime and target. The main result was that 6‐year‐old children showed both lexical and structural priming for both the active transitive and passive constructions, whereas 3‐ and 4‐year‐old children showed lexical priming only. This pattern of results would seem to indicate that 6‐year‐old children have relatively abstract representations of these constructions, whereas 3‐ and 4‐year‐old children have as an integral part of their representations certain specific lexical items, especially pronouns and some grammatical morphemes. In a second study it was found that children did not need to repeat the prime out loud in order to be primed – suggesting that the priming effect observed concerns not just peripheral production mechanisms but underlying linguistic representations common to comprehension and production. These results support the view that young children develop abstract linguistic representations gradually during the preschool years.
Tracking individuals via object‐files: evidence from infants’ manual search
Authors: Feigenson Lisa , Carey Susan
Published: November 2003

In two experiments, a manual search task explored 12‐ to 14‐month‐old infants’ representations of small sets of objects. In this paradigm, patterns of searching revealed the number of objects infants represented as hidden in an opaque box. In Experiment 1, we obtained the set‐size signature of object‐file representations: infants succeeded at representing precisely 1, precisely 2, and precisely 3 objects in the box, but failed at representing 4 (or even that 4 is greater than 2). In Experiment 2, we showed that infants’ expectations about the contents of the box were based on number of individual objects, and not on a continuous property such as total object volume. These findings support the hypothesis that infants maintained representations of individuals, that object‐files were the underlying means of representing these individuals, and that object‐file models can be compared via one‐to‐one correspondence to establish numerical equivalence.
Regime Types and Discrimination against Ethnoreligious Minorities: A Cross-Sectional Analysis of the Autocracy-Democracy Continuum
Authors: Fox Jonathan , Sandler Shmuel
Published: November 2003

Although many assume that the relationship between the autocracy-democracy continuum and discrimination is linear, with autocracies discriminating the most and democracies discriminating the least, the assumption is not universal. This study uses the Minorities at Risk dataset to test this relationship with regard to government treatment of religiously differentiated ethnic minorities (ethnoreligious minorities) as well as ethnic minorities that are not religiously differentiated. The results show that the pattern of treatment of ethnoreligious minorities differs from that of other ethnic minorities. The extent to which a state is democratic has no clear influence on the level of discrimination against non-religiously differentiated ethnic minorities, but it has a clear influence on the level of discrimination against ethnoreligious minorities. Autocracies discriminate more than democracies against ethnoreligious minorities, but semi-democracies, those governments that are situated between democracies and autocracies, discriminate even less. This result is consistent on all 11 measures used here and is statistically significant for seven of them, and it remains strong when controlling for other factors, including separatism. This phenomenon increases in strength from the beginning to the end of the 1990s. Also, democracies discriminate against ethnoreligious minorities more than they do against other minorities. The nature of liberal democracy may provide an explanation for this phenomenon.
Contractualism and Liberal Neutrality: A Defence
Authors: Lecce Steven A.
Published: November 2003

The most influential contemporary defences of liberal neutrality are premised on a contractual view of political legitimacy. For contractualists, perfectionist principles of justice are illegitimate because they cannot be the object of reasonable agreement among free and equal citizens. Several critics have challenged this connection between contractualism and neutrality by suggesting that the epistemic arguments commonly offered in its favour are self-defeating. This paper examines three recent expressions of this claim – those of Simon Caney, Simon Clarke and Joseph Chan – and finds that none of them succeeds. They fail because they mistake an ethical claim about how states should respond to disagreement for an epistemic one that explains why such a response is needed.
Against Restitution
Authors: Vernon Richard
Published: November 2003

Recently, states and other institutions have undertaken to make restitution for past abuses. Distinctions need to be made between various kinds of restitutive practices that rest on quite different normative grounds. Moreover, the core idea of restitution, in attaching obligation to particular historically grounded relationships, is questionable, and what is being attempted is better explained and justified in terms of a number of standard principles of justice of a non-restitutive kind; for although there is, in principle, a clear case of restitutive justice, its elements rarely, if ever, exist in the real world in an unmixed state. Although there are significant objections to deriving local obligations from principles of universal justice, they have no force in this case. Policies termed ‘restitutive’ may well be justifiable, but they are misdescribed.
Governing beyond the Centre: A Critique of the Anglo-Governance School
Authors: Marinetto Mike
Published: November 2003

One of the more intriguing theoretical discussions of recent years involves the concept of governance. There is now a substantial body of work concerning the way governance has affected the contribution of central government to the policy process. Possibly the most prominent and influential account of governance theory in British political science is offered by Rod Rhodes. His most recent writings have employed governance theory to explore the institutions, actors and processes of change within the core executive. His ‘Anglo-governance’ model has emerged as a prevalent and authoritative account of how new methods of governing have emerged in society. Significantly, it is maintained that a distinct shift has taken place in government, from a hierarchical organisation to a fragmented and decentralised entity that is heavily reliant on a range of complex and independent policy networks. There is undoubted evidence that government is a fractured institution that is dependent on state and non-state actors beyond the centre. This paper questions whether such features entail the emergence of a new form of governance. Central government is still highly resourced and has, at its disposal, a range of powers with which to retain influence over public sector agencies. Historical evidence also shows that the British polity has long been decentralised. Thus, it is difficult to see how recent developments have in any way transformed the capacities of the core executive. It seems that alternative ways of conceptualising the institutions, actors and processes of change in government are required. Recent efforts to develop ‘organising perspectives’, within the intellectual parameters of governance theory, offer a more ‘conceptually cautious’ treatment of the central state.
“Feminism Lite?” Feminist Identification, Speaker Appearance, and Perceptions of Feminist and Antifeminist Messengers
Authors: Bullock Heather E. , Fernald Julian L.
Published: November 2003

Drawing on a communications model of persuasion ( Hovland, Janis, & Kelley, 1953 ), this study examined the effect of target appearance on feminists' and nonfeminists' perceptions of a speaker delivering a feminist or an antifeminist message. One hundred three college women watched one of four videotaped speeches that varied by content (profeminist vs. antifeminist) and target appearance (“feminine” vs. “masculine”). Self-identified feminists responded more favorably to a profeminist message when it was presented by a feminine than a masculine-appearing speaker and expressed less feminist attitudes after viewing a masculine-appearing feminist speaker. Nonfeminists' evaluations did not vary as a function of speaker appearance or message content. Implications for the communication of feminist ideology are discussed.
Suspended Animation in
C. elegans
Requires the Spindle Checkpoint
Authors: Nystul Todd G. , Goldmark Jesse P. , Padilla Pamela A. , Roth Mark B.
Published: November 2003

In response to environmental signals such as anoxia, many organisms enter a state of suspended animation, an extreme form of quiescence in which microscopically visible movement ceases. We have identified a gene,
san-1
, that is required for suspended animation in
Caenorhabditis elegans
embryos. We show that
san-1
functions as a spindle checkpoint component in
C. elegans
. During anoxia-induced suspended animation, embryos lacking functional SAN-1 or a second spindle checkpoint component, MDF-2, failed to arrest the cell cycle, exhibited chromosome missegregation, and showed reduced viability. These data provide a model for how a dynamic biological process is arrested in suspended animation.
Increased Longevities of Post-Paleozoic Marine Genera After Mass Extinctions
Authors: Miller Arnold I. , Foote Michael
Published: November 2003

Cohorts of marine taxa that originated during recoveries from mass extinctions were commonly more widespread spatially than those originating at other times. Coupled with the recognition of a correlation between the geographic ranges and temporal longevities of marine taxa, this observation predicts that recovery taxa were unusually long-lived geologically. We analyzed this possibility by assessing the longevities of marine genus cohorts that originated in successive substages throughout the Phanerozoic. Results confirm that several mass extinction recovery cohorts were significantly longer lived than other cohorts, but this effect was limited to the post-Paleozoic, suggesting differences in the dynamics of Paleozoic versus post-Paleozoic diversification.
Control of Effector CD8
+
T Cell Function by the Transcription Factor
Eomesodermin
Authors: Pearce Erika L. , Mullen Alan C. , Martins Gislâine A. , Krawczyk Connie M. , Hutchins Anne S. , Zediak Valerie P. , Banica Monica , DiCioccio Catherine B. , Gross Darrick A. , Mao Chai-an , Shen Hao , Cereb Nezih , Yang Soo Y. , Lindsten Tullia , Rossant Janet , Hunter Christopher A. , Reiner Steven L.
Published: November 2003

Activated CD8
+
T cells play a critical role in host defense against viruses, intracellular microbes, and tumors. It is not clear if a key regulatory transcription factor unites the effector functions of CD8
+
T cells. We now show that
Eomesodermin
(
Eomes
), a paralogue of
T-bet
, is induced in effector CD8
+
T cells in vitro and in vivo. Ectopic expression of Eomes was sufficient to invoke attributes of effector CD8
+
T cells, including interferon-γ (IFN-γ), perforin, and granzyme B. Loss-of-function analysis suggests Eomes may also be necessary for full effector differentiation of CD8
+
T cells. We suggest that
Eomesodermin
is likely to complement the actions of
T-bet
and act as a key regulatory gene in the development of cell-mediated immunity.
Development and Validation of a Nested-PCR-Denaturing Gradient Gel Electrophoresis Method for Taxonomic Characterization of Bifidobacterial Communities
Authors: Temmerman R. , Masco L. , Vanhoutte T. , Huys G. , Swings J.
Published: November 2003

The taxonomic characterization of a bacterial community is difficult to combine with the monitoring of its temporal changes. None of the currently available identification techniques are able to visualize a “complete” community, whereas techniques designed for analyzing bacterial ecosystems generally display limited or labor-intensive identification potential. This paper describes the optimization and validation of a nested-PCR-denaturing gradient gel electrophoresis (DGGE) approach for the species-specific analysis of bifidobacterial communities from any ecosystem. The method comprises aBifidobacterium-specific PCR step, followed by purification of the amplicons that serve as template DNA in a second PCR step that amplifies the V3 and V6-V8 regions of the 16S rRNA gene. A mix of both amplicons is analyzed on a DGGE gel, after which the band positions are compared with a previously constructed database of reference strains. The method was validated through the analysis of four artificial mixtures, mimicking the possible bifidobacterial microbiota of the human and chicken intestine, a rumen, and the environment, and of two fecal samples. Except for the speciesBifidobacterium coryneformeandB. indicum, all currently known bifidobacteria originating from various ecosystems can be identified in a highly reproducible manner. Because no further cloning and sequencing of the DGGE bands is necessary, this nested-PCR-DGGE technique can be completed within a 24-h span, allowing the species-specific monitoring of temporal changes in the bifidobacterial community.
Rapid Identification of
Listeria
Species by Using Restriction Fragment Length Polymorphism of PCR-Amplified 23S rRNA Gene Fragments
Authors: Paillard Delphine , Dubois Véronique , Duran Robert , Nathier Fany , Guittet Catherine , Caumette Pierre , Quentin Claudine
Published: November 2003

A molecular method based on restriction fragment length polymorphism (RFLP) of PCR-amplified fragments of the 23S rRNA gene was designed to rapidly identify
Listeria
strains to the species level. Two fragments (S1, 460 bp, and S2, 890 bp) were amplified from boiled DNA. S2 was cut with the restriction enzymes
Xmn
I or
Cfo
I and, if needed, S1 was digested by either
Alu
I or
Cla
I. This method was first optimized with six reference strains and then applied to 182 isolates collected from effluents of treatment plants. All isolates were also identified by the API
Listeria
kit, hemolysis, and phosphatidylinositol-specific phospholipase C production (PI-PLC) on ALOA medium. The PCR-RFLP method unambiguously identified 160 environmental strains, including 131 in concordance with the API system, and revealed that 22 isolates were mixed cultures of
Listeria monocytogenes
and
Listeria innocua
. Discrepant results were resolved by a multiplex PCR on the
iap
gene, which confirmed the PCR-RFLP data for 49 of the 51 discordances, including the 22 mixed cultures. Sequencing of the 16S rRNA gene for 12 selected strains and reconstruction of a phylogenetic tree validated the molecular methods, except for two unclassifiable strains. The 158 single identifiable isolates were 92
L. monocytogenes
(including seven nonhemolytic and PI-PLC-negative strains), 61
L. innocua
, 4
Listeria seeligeri
, and 1
Listeria welshimeri
strain. The PCR-RFLP method proposed here provides rapid, easy-to-use, inexpensive, and reliable identification of the six
Listeria
species. Moreover, it can detect mixtures of
Listeria
species and thus is particularly adapted to environmental and food microbiology.
Assessment of Photodynamic Destruction of
Escherichia coli
O157:H7 and
Listeria monocytogenes
by Using ATP Bioluminescence
Authors: Romanova N. A. , Brovko L. Y. , Moore L. , Pometun E. , Savitsky A. P. , Ugarova N. N. , Griffiths M. W.
Published: November 2003

Antimicrobial photodynamic therapy was shown to be effective against a wide range of bacterial cells, as well as for fungi, yeasts, and viruses. It was shown previously that photodestruction of yeast cells treated with photosensitizers resulted in cell destruction and leakage of ATP. Three photosensitizers were used in this study: tetra(
N
-methyl-4-pyridyl)porphine tetratosylate salt (TMPyP), toluidine blue O (TBO), and methylene blue trihydrate (MB). A microdilution method was used to determine MICs of the photosensitizers against both
Escherichia coli
O157:H7 and
Listeria monocytogenes
. To evaluate the effects of photodestruction on
E. coli
and
L. monocytogenes
cells, a bioluminescence method for detection of ATP leakage and a colony-forming assay were used. All tested photosensitizers were effective for photodynamic destruction of both bacteria. The effectiveness of photosensitizers (in microgram-per-milliliter equivalents) decreased in the order TBO > MB > TMPyP for both organisms. The MICs were two- to fourfold higher for
E. coli
O157:H7 than for
L. monocytogenes
. The primary effects of all of the photosensitizers tested on live bacterial cells were a decrease in intracellular ATP and an increase in extracellular ATP, accompanied by elimination of viable cells from the sample. The time courses of photodestruction and intracellular ATP leakage were different for
E. coli
and
L. monocytogenes
. These results show that bioluminescent ATP-metry can be used for investigation of the first stages of bacterial photodestruction.
Innovative Approach for Improvement of an Antibiotic-Overproducing Industrial Strain of
Streptomyces albus
Authors: Tamehiro Norimasa , Hosaka Takeshi , Xu Jun , Hu Haifeng , Otake Noboru , Ochi Kozo
Published: November 2003

Working with a
Streptomyces albus
strain that had previously been bred to produce industrial amounts (10 mg/ml) of salinomycin, we demonstrated the efficacy of introducing drug resistance-producing mutations for further strain improvement. Mutants with enhanced salinomycin production were detected at a high incidence (7 to 12%) among spontaneous isolates resistant to streptomycin (Str
r
), gentamicin, or rifampin (Rif
r
). Finally, we successfully demonstrated improvement of the salinomycin productivity of the industrial strain by 2.3-fold by introducing a triple mutation. The Str
r
mutant was shown to have a point mutation within the
rpsL
gene (encoding ribosomal protein S12). Likewise, the Rif
r
mutant possessed a mutation in the
rpoB
gene (encoding the RNA polymerase β subunit). Increased productivity of salinomycin in the Str
r
mutant (containing the K88R mutation in the S12 protein) may be a result of an aberrant protein synthesis mechanism. This aberration may manifest itself as enhanced translation activity in stationary-phase cells, as we have observed with the poly(U)-directed cell-free translation system. The K88R mutant ribosome was characterized by increased 70S complex stability in low Mg
2+
concentrations. We conclude that this aberrant protein synthesis ability in the Str
r
mutant, which is a result of increased stability of the 70S complex, is responsible for the remarkable salinomycin production enhancement obtained.
Nitrogen-Regulated Hypermutator Strain of
Synechococcus
sp. for Use in In Vivo Artificial Evolution
Authors: Emlyn-Jones Daniel , Price G. Dean , Andrews T. John
Published: November 2003

Artificially evolved variants of proteins with roles in photosynthesis may be selected most conveniently by using a photosynthetic organism, such as a cyanobacterium, whose growth depends on the function of the target protein. However, the limited transformation efficiency of even the most transformable cyanobacteria wastes much of the diversity of mutant libraries of genes produced in vitro, impairing the coverage of sequence space. This highlights the advantages of an in vivo approach for generating diversity in the selection organism itself. We constructed two different hypermutator strains of
Synechococcus
sp. strain PCC 7942 by insertionally inactivating or nutritionally repressing the DNA mismatch repair gene,
mutS
. Inactivation of
mutS
greatly increases the mutation rate of the cyanobacterium's genes, leading to an up-to-300-fold increase in the frequency of resistance to the antibiotics rifampin and spectinomycin. In order to control the rate of mutation and to limit cellular damage resulting from prolonged hypermutation, we placed the uninterrupted
mutS
gene in the cyanobacterial chromosome under the transcriptional control of the cyanobacterial
nirA
promoter, which is repressed in the presence of NH
4
+
as an N source and derepressed in its absence. By removing or adding this substrate, hypermutation was activated or repressed as required. As expected, hypermutation caused by repression in P
nir
A-
mut
S transformants led to an accumulation of spectinomycin resistance mutations during growth.
Glycolytic Breakdown of Sulfoquinovose in Bacteria: a Missing Link in the Sulfur Cycle
Authors: Roy Alexander B. , Hewlins Michael J. E. , Ellis Andrew J. , Harwood John L. , White Graham F.
Published: November 2003

Sulfoquinovose (6-deoxy-6-sulfo-d-glucopyranose), formed by the hydrolysis of the plant sulfolipid, is a major component of the biological sulfur cycle. However, pathways for its catabolism are poorly delineated. We examined the hypothesis that mineralization of sulfoquinovose to inorganic sulfate is initiated by reactions of the glycolytic and/or Entner-Doudoroff pathways in bacteria. Metabolites of [U-13C]sulfoquinovose were identified by13C-nuclear magnetic resonance (NMR) in strains ofKlebsiellaandAgrobacteriumpreviously isolated for their ability to utilize sulfoquinovose as a sole source of carbon and energy for growth, and cell extracts were analyzed for enzymes diagnostic for the respective pathways.Klebsiellasp. strain ABR11 grew rapidly on sulfoquinovose, with major accumulations of sulfopropandiol (2,3-dihydroxypropanesulfonate) but no detectable release of sulfate. Later, when sulfoquinovose was exhausted and growth was very slow, sulfopropandiol disappeared and inorganic sulfate and small amounts of sulfolactate (2-hydroxy-3-sulfopropionate) were formed. InAgrobacteriumsp. strain ABR2, growth and sulfoquinovose disappearance were again coincident, though slower than that inKlebsiellasp. Release of sulfate was still late but was faster than that inKlebsiellasp., and no metabolites were detected by13C-NMR. Extracts of both strains grown on sulfoquinovose contained phosphofructokinase activities that remained unchanged when fructose 6-phosphate was replaced in the assay mixture with either glucose 6-phosphate or sulfoquinovose. The results were consistent with the operation of the Embden-Meyerhoff-Parnas (glycolysis) pathway for catabolism of sulfoquinovose. Extracts ofKlebsiellabut notAgrobacteriumalso contained an NAD+-dependent sulfoquinovose dehydrogenase activity, indicating that the Entner-Doudoroff pathway might also contribute to catabolism of sulfoquinovose.
A New
Bacteroides
Conjugative Transposon That Carries an
ermB
Gene
Authors: Gupta Anamika , Vlamakis Hera , Shoemaker Nadja , Salyers Abigail A.
Published: November 2003

The erythromycin resistance gene
ermB
has been found in a variety of gram-positive bacteria. This gene has also been found in
Bacteroides
species but only in six recently isolated strains; thus, the gene seems to have entered this genus only recently. One of the six
Bacteroides ermB
-containing isolates, WH207, could transfer
ermB
to
Bacteroides thetaiotaomicron
strain BT4001 by conjugation. WH207 was identified as a
Bacteroides uniformis
strain based on the sequence of its 16S rRNA gene. Results of pulsed-field gel electrophoresis experiments demonstrated that the transferring element was normally integrated into the
Bacteroides
chromosome. The element was estimated from pulsed-field gel data to be about 100 kb in size. Since the element appeared to be a conjugative transposon (CTn), it was designated CTnBST. CTnBST was able to mobilize coresident plasmids and the circular form of the mobilizable transposon NBU1 to
Bacteroides
and
Escherichia coli
recipients. A 13-kb segment that contained
ermB
was cloned and sequenced. Most of the open reading frames in this region had little similarity at the amino acid sequence level to any proteins in the sequence databases, but a 1,723-bp DNA segment that included a 950-bp segment downstream of
ermB
had a DNA sequence that was virtually identical to that of a segment of DNA found previously in a
Clostridium perfringens
strain. This finding, together with the finding that
ermB
is located on a CTn, supports the hypothesis that CTnBST could have entered
Bacteroides
from some other genus, possibly from gram-positive bacteria. Moreover, this finding supports the hypothesis that many transmissible antibiotic resistance genes in
Bacteroides
are carried on CTns.
Low-Temperature Isolation of Disease-Suppressive Bacteria and Characterization of a Distinctive Group of Pseudomonads
Authors: Johansson P. Maria , Wright Sandra A. I.
Published: November 2003

The influence of environmental factors during isolation on the composition of potential biocontrol isolates is largely unknown. Bacterial isolates that efficiently suppressed wheat seedling blight caused by
Fusarium culmorum
were found by isolating psychrotrophic, root-associated bacteria and by screening them in a bioassay that mimicked field conditions. The impact of individual isolation factors on the disease-suppressive index (DSI) of almost 600 isolates was analyzed. The bacteria originated from 135 samples from 62 sites in Sweden and Switzerland. The isolation factors that increased the probability of finding isolates with high DSIs were sampling from arable land, Swiss origin of samples, and origination of isolates from plants belonging to the family Brassicaceae. The colony morphology of the isolates was characterized and compared to DSIs, which led to identification of a uniform morphological group containing 57 highly disease-suppressive isolates. Isolates in this group were identified as
Pseudomonas
sp.; they were fluorescent on King's medium B and had characteristic crystalline structures in their colonies. These isolates were morphologically similar to seven strains that had previously been selected for suppression of barley net blotch caused by
Drechslera teres
. Members of this morphological group grow at 1.5°C and produce an antifungal polyketide (2,3-deepoxy-2,3-didehydrorhizoxin [DDR]). They have similar two-dimensional polyacrylamide gel electrophoresis protein profiles, phenotypic characteristics, and in vitro inhibition spectra of pathogens. In summary, in this paper we describe some isolation factors that are important for obtaining disease-suppressive bacteria in our system, and we describe a novel group of biocontrol pseudomonads.
Methicillin (Oxacillin)-Resistant
Staphylococcus aureus
Strains Isolated from Major Food Animals and Their Potential Transmission to Humans
Authors: Lee John Hwa
Published: November 2003

From May 2001 to April 2003, various types of specimens from cattle, pigs, and chickens were collected and examined for the presence of methicillin (oxacillin)-resistant
Staphylococcus aureus
(MRSA).
S. aureus
was isolated and positively identified by using Gram staining, colony morphology, tests for coagulase and urease activities, and an API Staph Ident system. Among 1,913 specimens collected from the animals, 421 contained
S. aureus
; of these, 28 contained
S. aureus
resistant to concentrations of oxacillin higher than 2 μg/ml. Isolates from 15 of the 28 specimens were positive by PCR for the
mecA
gene. Of the 15
mecA
-positive MRSA isolates, 12 were from dairy cows and 3 were from chickens. Antimicrobial susceptibility tests of
mecA
-positive MRSA strains were performed by the disk diffusion method. All isolates were resistant to members of the penicillin family, such as ampicillin, oxacillin, and penicillin. All isolates were also susceptible to amikacin, vancomycin, and trimethoprim-sulfamethoxazole. To determine molecular epidemiological relatedness of these 15 animal MRSA isolates to isolates from humans, random amplified polymorphic DNA (RAPD) patterns were generated by arbitrarily primed PCR. The RAPD patterns of six of the isolates from animals were identical to the patterns of certain isolates from humans. The antibiotypes of the six animal isolates revealed types similar to those of the human isolates. These data suggested that the genomes of the six animal MRSA isolates were very closely related to those of some human MRSA isolates and were a possible source of human infections caused by consuming contaminated food products made from these animals.
Construction of
Escherichia coli
Strains for Conversion of Nitroacetophenones to
ortho
-Aminophenols
Authors: Kadiyala Venkateswarlu , Nadeau Lloyd J. , Spain Jim C.
Published: November 2003

The predominant bacterial pathway for nitrobenzene (NB) degradation uses an NB nitroreductase and hydroxylaminobenzene (HAB) mutase to form the ring-fission substrate
ortho
-aminophenol. We tested the hypothesis that constructed strains might accumulate the aminophenols from nitroacetophenones and other nitroaromatic compounds. We constructed a recombinant plasmid carrying NB nitroreductase (
nbzA
) and HAB mutase A (
habA
) genes, both from
Pseudomonas pseudoalcaligenes
JS45, and expressed the enzymes in
Escherichia coli
JS995. IPTG (isopropyl-β-
d
-thiogalactopyranoside)-induced cells of strain JS995 rapidly and stoichiometrically converted NB to 2-aminophenol, 2-nitroacetophenone (2NAP) to 2-amino-3-hydroxyacetophenone (2AHAP), and 3-nitroacetophenone (3NAP) to 3-amino-2-hydroxyacetophenone (3AHAP). We constructed another recombinant plasmid containing the nitroreductase gene (
nfs1
) from
Enterobacter cloacae
and
habA
from strain JS45 and expressed the enzymes in
E. coli
JS996. Strain JS996 converted NB to 2-aminophenol, 2-nitrotoluene to 2-amino-3-methylphenol, 3-nitrotoluene to 2-amino-4-methylphenol, 4-nitrobiphenyl ether to 4-amino-5-phenoxyphenol, and 1-nitronaphthalene to 2-amino-1-naphthol. In larger-scale biotransformations catalyzed by strain JS995, 75% of the 2NAP transformed was converted to 2AHAP, whereas 3AHAP was produced stoichiometrically from 3NAP. The final yields of the aminophenols after extraction and recovery were >64%. The biocatalytic synthesis of
ortho
-aminophenols from nitroacetophenones suggests that strain JS995 may be useful in the biocatalytic production of a variety of substituted
ortho
-aminophenols from the corresponding nitroaromatic compounds.
Population Structure of
Alexandrium
(Dinophyceae) Cyst Formation-Promoting Bacteria in Hiroshima Bay, Japan
Authors: Adachi Masao , Kanno Tsukasa , Okamoto Ryo , Itakura Shigeru , Yamaguchi Mineo , Nishijima Toshitaka
Published: November 2003

A total of 31 bacterial isolates that have potential
Alexandrium
cyst formation-promoting activity (Alex-CFPB) were isolated from Hiroshima Bay (Japan), which is characterized by seasonal blooms of the toxic dinoflagellate
Alexandrium tamarense
. The population structure of Alex-CFPB was analyzed by means of restriction fragment length polymorphism analysis of the 16S rRNA genes (16S rDNA). Fourteen ribotypes, A to N, were observed among the 31 isolates of Alex-CFPB by using four restriction enzymes,
Mbo
I,
Hha
I,
Rsa
I and
Bst
UI. Among them, seven isolates, which were obtained from the seawater samples taken during the peak and termination periods of the
A. tamarense
bloom in 1998, belonged to ribotype A. This result suggests that bacterial strains of ribotype A may be dominant in the Alex-CFPB assemblages during these periods. The partial 16S rDNA-based phylogenetic tree of 10 ribotypes studied showed that nine of them fell into the
Rhodobacter
group of the α subclass of the
Proteobacteria
. Eight of nine ribotypes of the
Rhodobacter
group fell into the lineage of the
Roseobacter
subgroup, and one fell into the
Rhodobacter
subgroup. The non-
Rhodobacter
group type fell into the
Marinobacterium
-
Neptunomonas
-
Pseudomonas
group of the γ-
Proteobacteria
. Isolates of Alex-CFPB ribotypes A and C do not have clear growth-promoting activities but have strong cyst formation-promoting activities (CFPAs) under our laboratory conditions. These results show that the Alex-CFPB assemblage may consist of various bacteria that belong mainly to the
Roseobacter
group and have strong CFPAs. These results suggest that not only the
Alexandrium
cyst formation-inhibiting bacteria (Alex-CFIB) reported previously but also Alex-CFPB, especially bacteria of ribotype A, may play significant roles in the process of encystment and bloom dynamics of
Alexandrium
in the natural environment.
Diversity and Structure of Bacterial Communitiesin Arctic versus Antarctic PackIce
Authors: Brinkmeyer Robin , Knittel Katrin , Jürgens Jutta , Weyland Horst , Amann Rudolf , Helmke Elisabeth
Published: November 2003

A
comprehensive assessment of bacterial diversity and community
composition in arctic and antarctic pack ice was conducted through
cultivation and cultivation-independent molecular techniques. We
sequenced 16S rRNA genes from 115 and 87 pure cultures of bacteria
isolated from arctic and antarctic pack ice, respectively. Most of the
33 arctic phylotypes were >97% identical to previously
described antarctic species or to our own antarctic isolates. At both
poles, the α- and γ-proteobacteria and the
Cytophaga
-
Flavobacterium
group were the dominant
taxonomic bacterial groups identified by cultivation as well as by
molecular methods. The analysis of 16S rRNA gene clone libraries from
multiple arctic and antarctic pack ice samples revealed a high
incidence of closely overlapping 16S rRNA gene clone and isolate
sequences. Simultaneous analysis of environmental samples with
fluorescence in situ hybridization (FISH) showed that∼
95% of 4′,6′-diamidino-2-phenylindole
(DAPI)-stained cells hybridized with the general bacterial probe
EUB338. More than 90% of those were further assignable.
Approximately 50 and 36% were identified asγ
-proteobacteria in arctic and antarctic samples,respectively. Approximately 25% were identified asα
-proteobacteria, and 25% were identified as belonging
to the
Cytophaga
-
Flavobacterium
group. For the
quantification of specific members of the sea ice community, new
oligonucleotide probes were developed which target the genera
Octadecabacter
,
Glaciecola
,
Psychrobacter
,
Marinobacter
,
Shewanella
, and
Polaribacter
.
High FISH detection rates of these groups as well as high viable counts
corroborated the overlap of clone and isolate sequences. A terrestrial
influence on the arctic pack ice community was suggested by the
presence of limnic
phylotypes.
Physicochemical Conditions and Microbial Activities in the Highly Alkaline Gut of the Humus-Feeding Larva of
Pachnoda ephippiata
(Coleoptera: Scarabaeidae)
Authors: Lemke Thorsten , Stingl Ulrich , Egert Markus , Friedrich Michael W. , Brune Andreas
Published: November 2003

The soil macrofauna plays an important role in the carbon and nitrogen cycle of terrestrial ecosystems. In order to gain more insight into the role of the intestinal microbiota in transformation and mineralization of organic matter during gut passage, we characterized the physicochemical conditions, microbial activities, and community structure in the gut of our model organism, the humus-feeding larva of the cetoniid beetle
Pachnoda ephippiata
. Microsensor measurements revealed an extreme alkalinity in the midgut, with highest values (pH > 10) between the second and third crown of midgut ceca. Both midgut and hindgut were largely anoxic, but despite the high pH, the redox potential of the midgut content was surprisingly high even in the largest instar. However, reducing conditions prevailed in the hindgut paunch of all instars (E
h
∼ −100 mV). Both gut compartments possessed a pronounced gut microbiota, with highest numbers in the hindgut, and microbial fermentation products were present in high concentrations. The stimulation of hindgut methanogenesis by exogenous electron donors, such as H
2
, formate, and methanol, together with considerable concentrations of formate in midgut and hemolymph, suggests that midgut fermentations are coupled to methanogenesis in the hindgut by an intercompartmental transfer of reducing equivalents via the hemolymph. The results of a cultivation-based enumeration of the major metabolic groups in midgut and hindgut, which yielded high titers of lactogenic, propionigenic, and acetogenic bacteria, are in good agreement not only with the accumulation of microbial fermentation products in the respective compartments but also with the results of a cultivation-independent characterization of the bacterial communities reported in the companion paper (M. Egert, B. Wagner, T. Lemke, A. Brune, and M. W. Friedrich, Appl. Environ. Microbiol. 69:6659-6668, 2003).
Microbial Community Structure in Midgut and Hindgut of the Humus-Feeding Larva of
Pachnoda ephippiata
(Coleoptera: Scarabaeidae)
Authors: Egert Markus , Wagner Bianca , Lemke Thorsten , Brune Andreas , Friedrich Michael W.
Published: November 2003

The guts of soil-feeding macroinvertebrates contain a complex microbial community that is involved in the transformation of ingested soil organic matter. In a companion paper (T. Lemke, U. Stingl, M. Egert, M. W. Friedrich, and A. Brune, Appl. Environ. Microbiol. 69:6650-6658, 2003), we show that the gut of our model organism, the humivorous larva of the cetoniid beetle
Pachnoda ephippiata
, is characterized by strong midgut alkalinity, high concentrations of microbial fermentation products, and the presence of a diverse, yet unstudied microbial community. Here, we report on the community structure of bacteria and archaea in the midgut, hindgut, and food soil of
P. ephippiata
larvae, determined with cultivation-independent techniques. Clone libraries and terminal restriction fragment length polymorphism analysis of 16S rRNA genes revealed that the intestines of
P. ephippiata
larvae contain a complex gut microbiota that differs markedly between midgut and hindgut and that is clearly distinct from the microbiota in the food soil. The bacterial community is dominated by phylogenetic groups with a fermentative metabolism (
Lactobacillales
,
Clostridiales
,
Bacillales
, and
Cytophaga-Flavobacterium-Bacteroides
[CFB] phylum), which is corroborated by high lactate and acetate concentrations in the midgut and hindgut and by the large numbers of lactogenic and acetogenic bacteria in both gut compartments reported in the companion paper. Based on 16S rRNA gene frequencies,
Actinobacteria
dominate the alkaline midgut, while the hindgut is dominated by members of the CFB phylum. The archaeal community, however, is less diverse. 16S rRNA genes affiliated with mesophilic
Crenarchaeota
, probably stemming from the ingested soil, were most frequent in the midgut, whereas
Methanobacteriaceae
-related 16S rRNA genes were most frequent in the hindgut. These findings agree with the reported restriction of methanogenesis to the hindgut of
Pachnoda
larvae.
Induction and Resuscitation of Viable but Nonculturable
Salmonella enterica
Serovar Typhimurium DT104†
Authors: Gupte A. R. , de Rezende C. L. E. , Joseph S. W.
Published: November 2003

Salmonella enterica
serovar Typhimurium DT104 11601was tested for its ability to maintain viability in minimal, chemically defined solutions. Periodic monitoring of growth and survival in microcosms of different ion concentrations, maintained at various temperatures, showed a gradual decline in culturable organisms (∼235 days) at 5°C. Organisms maintained at a higher temperature (21°C) showed continuous, equivalent CFU per milliliter (∼10
6
) up to 400 days after inoculation. Fluorescence microscopy with Baclight revealed that nonculturable cells were actually viable, while observations with scanning electron microscopy showed that the cells had retained their structural integrity. Temperature upshift (56°C ± 0.5, 15 s) of the nonculturable organisms (5°C) in Trypticase soy broth followed by immediate inoculation onto Trypticase soy agar (TSA) gave evidence of resuscitation. Interestingly,
S. enterica
serovar Typhimurium DT104 from the microcosms at either 5°C (1 to 200 days) or 21°C (1 to 250 days) did not show enhanced growth after intermittent inoculation onto catalase-supplemented TSA. Furthermore, cells from 21°C microcosms exposed to oxidative and osmotic stress showed greater resistance to stresses over increasing times of exposure than did recently grown cells. It is possible that the exceptional survivability and resilience of this particular strain may in part reflect the growing importance of this multidrug-resistant organism, in general, as a cause of intestinal disease in humans. The fact that
S. enterica
serovar Typhimurium DT104 11601 is capable of modifying its physiological characteristics, including entry into and recovery from the viable but nonculturable state, suggests the overall possibility that
S. enterica
serovar Typhimurium DT104 may be able to respond uniquely to various adverse environmental conditions.
Biosynthesis of Yersiniabactin, a Complex Polyketide-Nonribosomal Peptide, Using
Escherichia coli
as a Heterologous Host
Authors: Pfeifer Blaine A. , Wang Clay C. C. , Walsh Christopher T. , Khosla Chaitan
Published: November 2003

The medicinal value associated with complex polyketide and nonribosomal peptide natural products has prompted biosynthetic schemes dependent upon heterologous microbial hosts. Here we report the successful biosynthesis of yersiniabactin (Ybt), a model polyketide-nonribosomal peptide hybrid natural product, using
Escherichia coli
as a heterologous host. After introducing the biochemical pathway for Ybt into
E. coli
, biosynthesis was initially monitored qualitatively by mass spectrometry. Next, production of Ybt was quantified in a high-cell-density fermentation environment with titers reaching 67 ± 21 (mean ± standard deviation) mg/liter and a volumetric productivity of 1.1 ± 0.3 mg/liter-h. This success has implications for basic and applied studies on Ybt biosynthesis and also, more generally, for future production of polyketide, nonribosomal peptide, and mixed polyketide-nonribosomal peptide natural products using
E. coli
.