bluuebunny/crossref_metadata_2025 huggingface dataset:
Content Validity and Nursing Sensitivity of Community‐Level Outcomes From the Nursing Outcomes Classification (NOC)
Authors: Head Barbara J. , Aquilino Mary Lober , Johnson Marion , Reed David , Maas Meridean , Moorhead Sue
Published: September 2004

Purpose

: To evaluate the content validity and nursing sensitivity of six community‐level outcomes from the Nursing Outcomes Classification (NOC; Johnson, Maas, & Moorhead, 2000).

Design and Methods

: A survey research design was used. Questionnaires were mailed to 300 public health nursing experts; 102 nurses responded. Experts evaluated between 11 and 30 indicators for each of the six outcomes for: (a) importance of the indicators for measuring the outcome, and (b) influence of nursing on the indicators. Content validity and nursing sensitivity of the outcomes were estimated with a modified Fehring technique.

Findings

: All outcomes were deemed important; only Community Competence had an outcome content validity score <.80. The outcome sensitivity score for Community Health: Immunity was .80; other outcome scores ranged from .62‐.70. Indicator ratios for all 102 indicators met the study criterion for importance, with 87% designated as critical and 13% as supplemental. Sensitivity ratios reflected judgments that 45% of the indicators were sensitive to nursing intervention.

Conclusions

: The study provided evidence of outcome content validity and nursing sensitivity of the study outcomes; further validation research is recommended, followed by testing of the study outcomes in clinical practice. Community‐level nursing‐sensitive outcomes will potentially enable study of the efficacy and effectiveness of public health interventions focused on improving health of populations and communities.
Development and Psychometric Evaluation of the Professional Practice Environment (PPE) Scale
Authors: Erickson Jeanette Ives , Duffy Mary E. , Gibbons M. Patricia , Fitzmaurice Joan , Ditomassi Marianne , Jones Dorothy
Published: September 2004

Purpose

: To describe the Professional Practice Environment (PPE) scale, its conceptual development and psychometric evaluation, and its uses in measuring eight characteristics of the professional practice environment in an acute care setting.

Design and Methods

: The 38‐item PPE Scale was validated on a sample of 849 professional practice staff at the Massachusetts General Hospital in Boston. Psychometric analysis included: item analysis, principal components analysis (PCA) with varimax rotation and Kaiser normalization, and internal consistency reliability using Cronbach's alpha coefficient.

Findings

: Eight components were shown, confirming the original conceptually derived model's structure and accounting for 61% of explained variance. Cronbach's alpha coefficients for the eight PPE subscales ranged from .78 to .88.

Conclusions

: Findings showed the 38‐item PPE Scale was reliable and valid for use in health outcomes research to examine the professional practice environment of staff working in acute care settings.
Towards ‘One‐Stop’ Cardiac MR Image Analysis
Authors: Lelieveldt B. P. F. , van der Geest R. J. , Reiber J. H. C.
Published: September 2004

Cardiac magnetic resonance imaging (CMRI) has evolved towards a powerful diagnostic imaging modality, enabling a detailed and quantitative assessment of the presence, extent and development of cardiovascular disease. Because of its versatility, CMRI can be regarded as a collection of ‘MR modalities’, each dedicated to different aspects of cardiac function: global function, regional function, valvular function, perfusion and coronary function; therefore, CMRI is increasingly referred to as the ‘one‐stop shop in cardiac imaging’. The increased need for quantification in clinical research and practice, combined with the high data volume of CMRI has generated a great demand for software tools that facilitate and further automate the accurate quantitative analysis of cardiovascular MR patient examinations. This paper highlights a few recent advances in automated post‐processing and quantification in the context of ‘one‐stop’ CMR patient examinations, and discusses them for the most common acquisition types.
Evolution of Coral Pigments Recreated
Authors: Ugalde Juan A. , Chang Belinda S. W. , Matz Mikhail V.
Published: September 2004

In proteins homologous to the green fluorescent protein (GFP), formation of red fluorescence requires three autocatalytic steps, whereas only two are needed for green fluorescence. Multiple red/green color diversification events in the GFP superfamily may reflect convergent evolution of the more complex three-step pathway. In the great star coral
Montastraea cavernosa
, a recreated common ancestor of green and red proteins turned out to be green, indicating that in this case red proteins evolved their color independently from most other homologous red proteins. Furthermore, red color appears to have evolved gradually by small incremental transitions.
Reverse Methanogenesis: Testing the Hypothesis with Environmental Genomics
Authors: Hallam Steven J. , Putnam Nik , Preston Christina M. , Detter John C. , Rokhsar Daniel , Richardson Paul M. , DeLong Edward F.
Published: September 2004

Microbial methane consumption in anoxic sediments significantly impacts the global environment by reducing the flux of greenhouse gases from ocean to atmosphere. Despite its significance, the biological mechanisms controlling anaerobic methane oxidation are not well characterized. One current model suggests that relatives of methane-producing
Archaea
developed the capacity to reverse methanogenesis and thereby to consume methane to produce cellular carbon and energy. We report here a test of the “reverse-methanogenesis” hypothesis by genomic analyses of methane-oxidizing
Archaea
from deep-sea sediments. Our results show that nearly all genes typically associated with methane production are present in one specific group of archaeal methanotrophs. These genome-based observations support previous hypotheses and provide an informed foundation for metabolic modeling of anaerobic methane oxidation.
The Emergence of Competition Between Model Protocells
Authors: Chen Irene A. , Roberts Richard W. , Szostak Jack W.
Published: September 2004

The transition from independent molecular entities to cellular structures with integrated behaviors was a crucial aspect of the origin of life. We show that simple physical principles can mediate a coordinated interaction between genome and compartment boundary, independent of any genomic functions beyond self-replication. RNA, encapsulated in fatty acid vesicles, exerts an osmotic pressure on the vesicle membrane that drives the uptake of additional membrane components, leading to membrane growth at the expense of relaxed vesicles, which shrink. Thus, more efficient RNA replication could cause faster cell growth, leading to the emergence of Darwinian evolution at the cellular level.
Electrically Driven Single-Cell Photonic Crystal Laser
Authors: Park Hong-Gyu , Kim Se-Heon , Kwon Soon-Hong , Ju Young-Gu , Yang Jin-Kyu , Baek Jong-Hwa , Kim Sung-Bock , Lee Yong-Hee
Published: September 2004

We report the experimental demonstration of an electrically driven, single-mode, low threshold current (∼260 μA) photonic band gap laser operating at room temperature. The electrical current pulse is injected through a sub-micrometer-sized semiconductor wire at the center of the mode with minimal degradation of the quality factor. The actual mode of interest operates in a nondegenerate monopole mode, as evidenced through the comparison of the measurement with the computation based on the actual fabricated structural parameters. As a small step toward a thresholdless laser or a single photon source, this wavelength-size photonic crystal laser may be of interest to photonic crystals, cavity quantum electrodynamics, and quantum information communities.
Aging in Rhesus Monkeys: Relevance to Human Health Interventions
Authors: Roth George S. , Mattison Julie A. , Ottinger Mary Ann , Chachich Mark E. , Lane Mark A. , Ingram Donald K.
Published: September 2004

Progress in gerontological research has been promoted through the use of numerous animal models, which have helped identify possible mechanisms of aging and age-related chronic diseases and evaluate possible interventions with potential relevance to human aging and disease. Further development of nonhuman primate models, particularly rhesus monkeys, could accelerate this progress, because their closer genetic relationship to humans produces a highly similar aging phenotype. Because the relatively long lives of primates increase the administrative and economic demands on research involving them, new emphasis has emerged on increasing the efficient use of these valuable resources through cooperative, interdisciplinary research.
Comparison of Diversities and Compositions of Bacterial Populations Inhabiting Natural Forest Soils
Authors: Hackl Evelyn , Zechmeister-Boltenstern Sophie , Bodrossy Levente , Sessitsch Angela
Published: September 2004

The diversity and composition of soil bacterial communities were compared among six Austrian natural forests, including oak-hornbeam, spruce-fir-beech, and Austrian pine forests, using terminal restriction fragment length polymorphism (T-RFLP, or TRF) analysis and sequence analysis of 16S rRNA genes. The forests studied differ greatly in soil chemical characteristics, microbial biomass, and nutrient turnover rates. The aim of this study was to relate these differences to the composition of the bacterial communities inhabiting the individual forest soils. Both TRF profiling and clone sequence analysis revealed that the bacterial communities in soils under Austrian pine forests, representing azonal forest types, were distinct from those in soils under zonal oak-hornbeam and spruce-fir-beech forests, which were more similar in community composition. Clones derived from an Austrian pine forest soil were mostly affiliated with high-G+C gram-positive bacteria (49%), followed by members of the α
-Proteobacteria
(20%) and the
Holophaga/Acidobacterium
group (12%). Clones in libraries from oak-hornbeam and spruce-fir-beech forest soils were mainly related to the
Holophaga/Acidobacterium
group (28 and 35%), followed by members of the
Verrucomicrobia
(24%) and the α
-Proteobacteria
(27%), respectively. The soil bacterial communities in forests with distinct vegetational and soil chemical properties appeared to be well differentiated based on 16S rRNA gene phylogeny. In particular, the outstanding position of the Austrian pine forests, which are determined by specific soil conditions, was reflected in the bacterial community composition.
Calicivirus Inactivation by Nonionizing (253.7-Nanometer-Wavelength [UV]) and Ionizing (Gamma) Radiation
Authors: de Roda Husman Ana Maria , Bijkerk Paul , Lodder Willemijn , van den Berg Harold , Pribil Walter , Cabaj Alexander , Gehringer Peter , Sommer Regina , Duizer Erwin
Published: September 2004

Noroviruses (previously Norwalk-like viruses) are the most common viral agents associated with food- and waterborne outbreaks of gastroenteritis. In the absence of culture methods for noroviruses, animal caliciviruses were used as model viruses to study inactivation by nonionizing (253.7-nm-wavelength [UV]) and ionizing (gamma) radiation. Here, we studied the respiratory feline calicivirus (FeCV) and the presumed enteric canine calicivirus (CaCV) and compared them with the well-studied bacteriophage MS2. When UV irradiation was used, a 3-log
10
reduction was observed at a fluence of 120 J/m
2
in the FeCV suspension and at a fluence of 200 J/m
2
for CaCV; for the more resistant phage MS2 there was a 3-log
10
reduction at a fluence of 650 J/m
2
. Few or no differences were observed between levels of UV inactivation in high- and low-protein-content virus stocks. In contrast, ionizing radiation could readily inactivate MS2 in water, and there was a 3-log
10
reduction at a dose of 100 Gy, although this did not occur when the phage was diluted in high-protein-content stocks of CaCV or FeCV. The low-protein-content stocks showed 3-log
10
reductions at a dose of 500 Gy for FeCV and at a dose of 300 for CaCV. The inactivation rates for both caliciviruses with ionizing and nonionizing radiation were comparable but different from the inactivation rates for MS2. Although most FeCV and CaCV characteristics, such as overall particle and genome size and structure, are similar, the capsid sequences differ significantly, making it difficult to predict human norovirus inactivation. Adequate management of UV and gamma radiation processes for virus inactivation should limit public health risks.
The
gnyRDBHAL
Cluster Is Involved in Acyclic Isoprenoid Degradation in
Pseudomonas aeruginosa
Authors: Díaz-Pérez A. L. , Zavala-Hernández A. N. , Cervantes C. , Campos-García J.
Published: September 2004

Pseudomonas aeruginosa
PAO1 mutants affected in the ability to degrade acyclic isoprenoids were isolated with transposon mutagenesis. The
gny
cluster (for geranoyl), which encodes the enzymes involved in the lower pathway of acyclic isoprenoid degradation, was identified. The
gny
cluster is constituted by five probable structural genes,
gnyDBHAL
, and a possible regulatory gene,
gnyR
. Mutations in the
gnyD
,
gnyB
,
gnyA
, or
gnyL
gene caused inability to assimilate acyclic isoprenoids of the citronellol family of compounds. Transcriptional analysis showed that expression of the
gnyB
gene was induced by citronellol and repressed by glucose, whereas expression of the
gnyR
gene had the opposite behavior. Western blot analysis of citronellol-grown cultures showed induction of biotinylated proteins of 70 and 73 kDa, which probably correspond to 3-methylcrotonoyl-coenzyme A (CoA) carboxylase and geranoyl-CoA carboxylase (GCCase) alpha subunits, respectively. The 73-kDa biotinylated protein, identified as the α-GCCase subunit, is encoded by
gnyA
. Intermediary metabolites of the isoprenoid pathway, citronellic and geranic acids, were shown to accumulate in
gnyB
and
gnyA
mutants. Our data suggest that the protein products encoded in the
gny
cluster are the β and α subunits of geranoyl-CoA carboxylase (GnyB and GnyA), the citronelloyl-CoA dehydrogenase (GnyD), the γ-carboxygeranoyl-CoA hydratase (GnyH), and the 3-hydroxy-γ-carboxygeranoyl-CoA lyase (GnyL). We conclude that the
gnyRDBHAL
cluster is involved in isoprenoid catabolism.
Comparison of ATPase-Encoding Type III Secretion SystemhrcNGenes in Biocontrol Fluorescent Pseudomonads and in Phytopathogenic Proteobacteria
Authors: Rezzonico Fabio , Défago Geneviève , Moënne-Loccoz Yvan
Published: September 2004

Type III protein secretion systems play a key role in the virulence of many pathogenic proteobacteria, but they also occur in nonpathogenic, plant-associated bacteria. Certain type III protein secretion genes (e.g.,hrcC) have been found inPseudomonassp. strain SBW25 (and other biocontrol pseudomonads), but other type III protein secretion genes, such as the ATPase-encoding genehrcN, have not been found. Using both colony hybridization and a PCR approach, we show here thathrcNis nevertheless present in many biocontrol fluorescent pseudomonads. The phylogeny of biocontrolPseudomonasstrains based on partialhrcNsequences was largely congruent with the phylogenies derived from analyses ofrrs(encoding 16S rRNA) and, to a lesser extent, biocontrol genes, such asphlD(for 2,4-diacetylphloroglucinol production) andhcnBC(for HCN production). Most biocontrol pseudomonads clustered separately from phytopathogenic proteobacteria, including pathogenic pseudomonads, in thehrcNtree. The exception was strain KD, which clustered with phytopathogenic pseudomonads, such asPseudomonas syringae, suggesting thathrcNwas acquired from the latter species. Indeed, strain KD (unlike strain SBW25) displayed the same organization of thehrpJoperon, which containshrcN, asP. syringae. These results indicate that the occurrence ofhrcNin most biocontrol pseudomonads is not the result of recent horizontal gene transfer from phytopathogenic bacteria, although such transfer might have occurred for a minority of biocontrol strains.
Clonality and Diversity of the Fish Pathogen
Lactococcus garvieae
in Mediterranean Countries
Authors: Eyngor Marina , Zlotkin Amir , Ghittino Claudio , Prearo Marino , Douet Diane-Gaëlle , Chilmonczyk Stefan , Eldar Avi
Published: September 2004

Infection with
Lactococcus garvieae
is considered the most important risk factor for the European trout industry, and the losses are approximately 50% of the total production. To improve our understanding of the genetic links among strains originating from different countries, we examined the population structure of
L. garvieae
by comparing 81 strains isolated from different sources and ecosystems (41 farms in six countries) in which the bacterium is commonly found. Genetic similarities (as assessed with molecular tools, including restriction fragment length polymorphism ribotyping with two endonucleases) were compared with serological data. The combined results reveal that in endemic sites the bacterial population displays a clonal structure, whereas bacterial diversity characterizes sites where the infection is sporadic.
Modulation of
Aspergillus awamori
Thaumatin Secretion by Modification of
bipA
Gene Expression
Authors: Lombraña Marta , Moralejo Francisco J. , Pinto Rosa , Martín Juan F.
Published: September 2004

Two different strains,
Aspergillus awamori
TGDTh-4 and
A. awamori
TGP-3 overexpressing a synthetic gene encoding the plant sweet protein thaumatin, showed an unfolded protein response. To facilitate protein secretion, the chaperone BiPA gene was expressed in
A. awamori
under control of the strong constitutive promoter of the
gpdA
gene. A good correlation was observed between the level of the
bipA
transcript in different strains and the amount of thaumatin secreted. Thaumatin secretion was increased 2- to 2.5-fold in transformants overexpressing the
bipA
gene compared with the parental strain. Secretion of the homologous proteins α-amylase and glucoamylase was not affected by the
bipA
gene overexpression. The requirement for BiPA for secretion of thaumatin was confirmed by attenuation of the endogenous
bipA
gene expression with an antisense RNA cassette. The decrease in
bipA
expression reduced the amount of secreted thaumatin up to 80% without affecting the secretion of the homologous α-amylase and glucoamylase proteins. The BiPA protein is, therefore, very important for secretion of some heterologous proteins, such as thaumatin in
A. awamori
.
Genotyping and Toxigenic Potential ofBacillus subtilisandBacillus pumilusStrains Occurring in Industrial and Artisanal Cured Sausages
Authors: Matarante Alessandra , Baruzzi Federico , Cocconcelli Pier Sandro , Morea Maria
Published: September 2004

Artisanal and industrial sausages were analyzed for their aerobic, heat-resistant microflora to assess whether new emerging pathogens could be present amongBacillusstrains naturally contaminating cured meat products. Sixty-four isolates were characterized by randomly amplified polymorphic DNA (RAPD)-PCR and fluorescent amplified fragment length polymorphism (fAFLP). The biotypes, identified by partial 16S rRNA gene sequence analysis, belonged toBacillus subtilis,Bacillus pumilus, andBacillus amyloliquefaciensspecies. Both RAPD-PCR and fAFLP analyses demonstrated that a high genetic heterogeneity is present in theB. subtilisgroup even in strains harvested from the same source, making it possible to isolate 56 different biotypes. Moreover, fAFLP analysis made it possible to distinguishB. subtilisfromB. pumilusstrains. The strains were characterized for their toxigenic potential by molecular, physiological, and immunological techniques. Specific PCR analyses revealed the absence of DNA sequences related to HBL, BcET, NHE, and entFMBacillus cereusenterotoxins and the enzymes sphingomyelinase Sph and phospholipase PI-PLC in all strains; also, the immunological analyses showed thatBacillusstrains did not react with NHE- and HBL-specific antibodies. However, some isolates were found to be positive for hemolytic and lecithinase activity. The absence of toxigenic potential inBacillusstrains from the sausages analyzed indicates that these products can be considered safe under the processing conditions they were produced; however, great care should be taken when the ripening time is shortened, particularly in the case of traditional sausages, which could contain high amounts ofBacillusstrains and possibly someB. cereuscells.
Identification of Specific In Vivo-Induced (ivi) Genes inYersinia ruckeriand Analysis of Ruckerbactin, a Catecholate Siderophore Iron Acquisition System
Authors: Fernández L. , Márquez I. , Guijarro J. A.
Published: September 2004

This work reports the utilization of an in vivo expression technology system to identify in vivo-induced (ivi) genes inYersinia ruckeriafter determination of the conditions needed for its selection in fish. Fourteen clones were selected, and the cloned DNA fragments were analyzed after partial sequencing. In addition to sequences with no significant similarity, homology with genes encoding proteins putatively involved in two-component and type IV secretion systems, adherence, specific metabolic functions, and others were found. Among these sequences, four were involved in iron acquisition through a catechol siderophore (ruckerbactin). Thus, unlike other pathogenic yersiniae producing yersiniabactin,Y. ruckerimight be able to produce and utilize only this phenolate. The genetic organization of the ruckerbactin biosynthetic and uptake loci was similar to that of theEscherichia colienterobactin gene cluster. GenesrucCandrupG, putative counterparts ofE. coli entCandfepG, respectively, involved in the biosynthesis and transport of the iron siderophore complex, respectively, were analyzed further. Thus, regulation of expression by iron and temperature and their presence in otherY. ruckerisiderophore-producing strains were confirmed for these two loci. Moreover, 50% lethal dose values 100-fold higher than those of the wild-type strain were obtained with therucCisogenic mutant, showing the importance of ruckerbactin in the pathogenesis caused by this microorganism.
In Situ Monitoring of Streptothricin Production byStreptomyces rocheiF20 in Soil and Rhizosphere
Authors: Anukool Usanee , Gaze William H. , Wellington Elizabeth M. H.
Published: September 2004

The onset of streptothricin (ST) biosynthesis inStreptomyces rocheiF20 was studied by using reverse transcription-PCR (RT-PCR) to detect transcripts of ST genes during growth in liquid medium, soil, and the rhizosphere. In situ results correlated with those obtained in vitro, illustrating the growth phase-dependent manner of ST production by F20. Maximal transcription of ST resistance (sttR) and biosynthesis (sttA) genes occurred during the transition between the exponential and stationary phases of growth, when the specific growth rate (μ) started to decline. A higher level of gene expression ofsttRversussttAwas observed in all experiments. In liquid culture, maximal transcript accumulation of thesttAgene was only ca. 40% that of thesttRgene.sttAandsttRmRNAs were detected in soil containing approximately 106CFU of growing cells g of soil−1.sttRmRNA was detected in sterile and nonsterile rhizosphere colonized with growing mycelium of F20 at 1.2 × 106and 4.0 × 105CFU g of soil−1, respectively. However, neithersttRnorsttAtranscripts were detected by RT-PCR in the rhizoplane, which supported a lower population density of F20 than the rhizosphere.
Anaerobic Growth of
Bacillus mojavensis
and
Bacillus subtilis
Requires Deoxyribonucleosides or DNA
Authors: Folmsbee Martha J. , McInerney Michael J. , Nagle David P.
Published: September 2004

Bacillus mojavensis
strains JF-2 (ATCC 39307), ROB2, and ABO21191
T
and
Bacillus subtilis
strains 168 (ATCC 23857) and ATCC 12332 required four deoxyribonucleosides or DNA for growth under strict anaerobic conditions.
Bacillus licheniformis
strains L89-11 and L87-11,
Bacillus sonorensis
strain TG8-8, and
Bacillus cereus
(ATCC 14579) did not require DNA for anaerobic growth. The requirement for the deoxyribonucleosides or DNA did not occur under aerobic growth conditions. The addition of a mixture of five nucleic acid bases, four ribonucleotides, or four ribonucleosides to the basal medium did not replace the requirement of
B. mojavensis
JF-2 for the four deoxyribonucleosides. However, the addition of salmon sperm DNA, herring sperm DNA,
Escherichia coli
DNA, or synthetic DNA (single or double stranded) to the basal medium supported anaerobic growth. The addition of four deoxyribonucleosides to the basal medium allowed aerobic growth of
B. mojavensis
JF-2 in the presence of hydroxyurea.
B. mojavensis
did not grow in DNA-supplemented basal medium that lacked sucrose as the energy source. These data provide strong evidence that externally supplied deoxyribonucleosides can be used to maintain a balanced deoxyribonucleotide pool for DNA synthesis and suggest that ribonucleotide reductases may not be essential to the bacterial cell cycle nor are they necessarily part of a minimal bacterial genome.
Degradation of Estrogens by
Rhodococcus zopfii
and
Rhodococcus equi
Isolates from Activated Sludge in Wastewater Treatment Plants
Authors: Yoshimoto Takeshi , Nagai Fumiko , Fujimoto Junji , Watanabe Koichi , Mizukoshi Harumi , Makino Takashi , Kimura Kazumasa , Saino Hideyuki , Sawada Haruji , Omura Hiroshi
Published: September 2004

We have isolated four strains of
Rhodococcus
which specifically degrade estrogens by using enrichment culture of activated sludge from wastewater treatment plants. Strain Y 50158, identified as
Rhodococcus zopfii
, completely and rapidly degraded 100 mg of 17β-estradiol, estrone, estriol, and ethinyl estradiol/liter, as demonstrated by thin-layer chromatography and gas chromatography-mass spectrometry analyses. Strains Y 50155, Y 50156, and Y 50157, identified as
Rhodococcus equi
, showed degradation activities comparable with that of Y 50158. Using the random amplified polymorphism DNA fingerprinting test, these three strains were confirmed to have been derived from different sources.
R. zopfii
Y 50158, which showed the highest activity among these four strains, revealed that the strain selectively degraded 17β-estradiol during jar fermentation, even when glucose was used as a readily utilizable carbon source in the culture medium. Measurement of estrogenic activities with human breast cancer-derived MVLN cells showed that these four strains each degraded 100 mg of 17β-estradiol/liter to 1/100 of the specific activity level after 24 h. It is thus suggested that these strains degrade 17β-estradiol into substances without estrogenic activity.
Transmission and Infectious Dose ofEscherichia coliO157:H7 in Swine
Authors: Cornick N. A. , Helgerson A. F.
Published: September 2004

Escherichia coliO157:H7 is only occasionally isolated from healthy swine, but some experimentally infected animals will shed the organism in their feces for at least 2 months. Potential explanations for the paucity of naturally occurring infections in swine, as compared to cattle, include a lack of animal-to-animal transmission so that the organism cannot be maintained within a herd, a high infectious dose, or herd management practices that prevent the maintenance of the organism in the gastrointestinal tract. We hypothesized that donor pigs infected withE. coliO157:H7 would transmit the organism to naïve pigs. We also determined the infectious dose and whether housing pigs individually on grated floors would decrease the magnitude or duration of fecal shedding. Infected donor pigs shedding <104CFU ofE. coliO157:H7 per g transmitted the organism to 6 of 12 naïve pigs exposed to them. The infectious dose ofE. coliO157:H7 for 3-month-old pigs was approximately 6 × 103CFU. There was no difference in the magnitude and duration of fecal shedding by pigs housed individually on grates compared to those housed two per pen on cement floors. These results suggest that swine do not have an innate resistance to colonization byE. coliO157:H7 and that they could serve as a reservoir host under suitable conditions.
Effect of Forage or Grain Diets with or without Monensin on Ruminal Persistence and Fecal
Escherichia coli
O157:H7 in Cattle
Authors: Van Baale M. J. , Sargeant J. M. , Gnad D. P. , DeBey B. M. , Lechtenberg K. F. , Nagaraja T. G.
Published: September 2004

Twelve ruminally cannulated cattle, adapted to forage or grain diet with or without monensin, were used to investigate the effects of diet and monensin on concentration and duration of ruminal persistence and fecal shedding of
E. coli
O157:H7. Cattle were ruminally inoculated with a strain of
E. coli
O157:H7 (10
10
CFU/animal) made resistant to nalidixic acid (Nal
r
). Ruminal and fecal samples were collected for 11 weeks, and then cattle were euthanized and necropsied and digesta from different gut locations were collected. Samples were cultured for detection and enumeration of Nal
r
E. coli
O157:H7. Cattle fed forage diets were culture positive for
E. coli
O157:H7 in the feces for longer duration (
P
< 0.05) than cattle fed a grain diet. In forage-fed cattle, the duration they remained culture positive for
E. coli
O157:H7 was shorter (
P
< 0.05) when the diet included monensin. Generally, ruminal persistence of Nal
r
E. coli
O157:H7 was not affected by diet or monensin. At necropsy,
E. coli
O157:H7 was detected in cecal and colonic digesta but not from the rumen. Our study showed that cattle fed a forage diet were culture positive longer and with higher numbers than cattle on a grain diet. Monensin supplementation decreased the duration of shedding with forage diet, and the cecum and colon were culture positive for
E. coli
O157:H7 more often than the rumen of cattle.
RsbT and RsbV Contribute to σB-Dependent Survival under Environmental, Energy, and Intracellular Stress Conditions inListeria monocytogenes
Authors: Chaturongakul Soraya , Boor Kathryn J.
Published: September 2004

Sigma B (σB) is a stress-responsive alternative sigma factor that has been identified in various gram-positive bacteria. Seven different regulators of sigma B (Rsbs) are located in thesigBoperons of bothBacillus subtilisandListeria monocytogenes. InB. subtilis, these proteins contribute to regulation of σBactivity by conveying environmental and energy stress signals through two well-established branches of a signal transduction pathway. RsbT contributes to regulation of σBactivity in response to environmental stresses, while RsbV contributes to σBactivation under both environmental and energy stresses inB. subtilis. To probeL. monocytogenesRsb roles in σB-mediated responses to various stresses, in-frame deletions were created inrsbTandrsbV. Phenotypic characterization of theL. monocytogenes rsbTandrsbVnull mutants revealed that both mutants were similar to the ΔsigBstrain in their abilities to survive under environmental stress conditions (exposure to synthetic gastric fluid, pH 2.5, acidified brain heart infusion broth [BHI], or oxidative stress [13 mM cumene hydroperoxide]). Under energy stress conditions (carbon starvation in defined media, entry into stationary phase, or reduced intracellular ATP), both ΔrsbTand ΔrsbVshowed survival reductions similar to that of the ΔsigBstrain. These observations suggest that the pathways for Rsb-dependent regulation of σBactivity differ betweenL. monocytogenesandB. subtilis. As σBalso activates transcription of theL. monocytogenes prfAP2 promoter, we evaluated virulence-associated characteristics of ΔprfAP1rsbTand ΔprfAP1rsbVdouble mutants in hemolysis and tissue culture assays. Both double mutants showed identical phenotypes to ΔprfAP1P2 and ΔprfAP1sigBdouble mutants, i.e., reduced hemolysis activity and reduced plaque size in mouse fibroblast cells. These findings indicate that RsbT and RsbV both contribute to σBactivation inL. monocytogenesduring exposure to environmental and energy stresses as well as during tissue culture infection.
Epiphytic Cyanobacteria onChara vulgarisAre the Main Contributors to N2Fixation in Rice Fields
Authors: Ariosa Yoanna , Quesada Antonio , Aburto Juan , Carrasco David , Carreres Ramón , Leganés Francisco , Fernández Valiente Eduardo
Published: September 2004

The distribution of nitrogenase activity in the rice-soil system and the possible contribution of epiphytic cyanobacteria on rice plants and other macrophytes to this activity were studied in two locations in the rice fields of Valencia, Spain, in two consecutive crop seasons. The largest proportion of photodependent N2fixation was associated with the macrophyteChara vulgarisin both years and at both locations. The nitrogen fixation rate associated withCharaalways represented more than 45% of the global nitrogenase activity measured in the rice field. The estimated average N2fixation rate associated withCharawas 27.53 kg of N ha−1crop−1. The mean estimated N2fixation rates for the other parts of the system for all sampling periods were as follows: soil, 4.07 kg of N ha−1crop−1; submerged parts of rice plants, 3.93 kg of N ha−1crop−1; and roots, 0.28 kg of N ha−1crop−1. Micrographic studies revealed the presence of epiphytic cyanobacteria on the surface ofChara. Three-dimensional reconstructions by confocal scanning laser microscopy revealed no cyanobacterial cells inside theCharastructures. Quantification of epiphytic cyanobacteria by image analysis revealed that cyanobacteria were more abundant in nodes than in internodes (on average, cyanobacteria covered 8.4% ± 4.4% and 6.2% ± 5.0% of the surface area in the nodes and internodes, respectively). Epiphytic cyanobacteria were also quantified by using a fluorometer. This made it possible to discriminate which algal groups were the source of chlorophylla. Chlorophyllameasurements confirmed that cyanobacteria were more abundant in nodes than in internodes (on average, the chlorophyllaconcentrations were 17.2 ± 28.0 and 4.0 ± 3.8 μg mg [dry weight] ofChara−1in the nodes and internodes, respectively). These results indicate that this macrophyte, which is usually considered a weed in the context of rice cultivation, may help maintain soil N fertility in the rice field ecosystem.
New Expression System Tightly Controlled by Zinc Availability in
Lactococcus lactis
Authors: Llull D. , Poquet I.
Published: September 2004

Here we developed the new expression system P
Zn
zitR
, based on the regulatory signals (P
Zn
promoter and
zitR
repressor) of the
Lactococcus lactis zit
operon, involved in Zn
2+
high-affinity uptake and regulation. A P
Zn
zitR
-controlled expression vector was constructed, and expression regulation was studied with two reporter genes,
uspnuc
and
lacLM
; these genes encode, respectively, a protein derived from
Staphylococcus aureus
secreted nuclease and
Leuconostoc mesenteroides
cytoplasmic β-galactosidase. Nuclease and β-galactosidase activities of
L. lactis
MG1363 cells expressing either
uspnuc
or
lacLM
under the control of P
Zn
zitR
were evaluated on plates and quantified from liquid cultures as a function of divalent metal ion, particularly Zn
2+
, availability in the environment. Our results demonstrate that P
Zn
zitR
is highly inducible upon divalent cation starvation, obtained either through EDTA addition or during growth in chemically defined medium, and is strongly repressed in the presence of excess Zn
2+
. The efficiency of the P
Zn
zitR
expression system was compared to that of the well-known nisin-controlled expression (NICE) system with the same reporter genes cloned under either P
Zn
zitR
or P

nisA
nisRK

control.
lacLM
induction levels reached with both systems were on the same order of magnitude, even though the NICE system is fivefold more efficient than the P
Zn
zitR
system. An even smaller difference or no difference was observed after 3 h of induction when nuclease was used as a reporter for Western blotting detection. P
Zn
zitR
proved to be a powerful expression system for
L. lactis
, as it is tightly controlled by the zinc concentration in the medium.
Geochemical Rate-RNA Integration Study: Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Gene Transcription and Photosynthetic Capacity of Planktonic Photoautotrophs
Authors: Corredor Jorge E. , Wawrik Boris , Paul John H. , Tran Hiep , Kerkhof Lee , López José M. , Dieppa Angel , Cárdenas Oswaldo
Published: September 2004

A pilot field experiment to assess the relationship between traditional biogeochemical rate measurements and transcriptional activity of microbial populations was carried out at the LEO 15 site off Tuckerton, N.J. Here, we report the relationship between photosynthetic capacity of autotrophic plankton and transcriptional activity of the large subunit gene (
rbcL
) for ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), the enzyme responsible for primary carbon fixation during photosynthesis. Similar diel patterns of carbon fixation and
rbcL
gene expression were observed in three of four time series, with maxima for photosynthetic capacity (
P
max
) and
rbcL
mRNA occurring between 10 a.m. and 1 p.m.. The lowest
P
max
and
rbcL
levels were detected between 6 p.m. and 10:30 p.m.. A significant correlation was found between
P
max
and form ID
rbcL
mRNA (
R
2
= 0.56) and forms IA and IB (
R
2
= 0.41 and 0.47, respectively). The correlation between the abundance of “diatom”
rbcL
and
P
max
mRNA was modest (
R
2
= 0.49;
n
= 12) but improved dramatically (
R
2
= 0.97;
n
= 10) upon removal of two outliers which represented afternoon samples with high
P
max
but lower mRNA levels. Clone libraries from reverse transcription-PCR-amplified
rbcL
mRNA indicated the presence of several chromophytic algae (diatoms, prymnesiophytes, and chrysophytes) and some eukaryotic green flagellates. Analogous results were obtained from amplified small rRNA sequences and secondary pigment analysis. These results suggest that diatoms were a major contributor to carbon fixation at LEO 15 at the time of sampling and that photosynthetic carbon fixation was partially controlled by transcriptional regulation of the RubisCO gene.
Effects of Temperature and Salinity on
Vibrio vulnificus
Population Dynamics as Assessed by Quantitative PCR
Authors: Randa Mark A. , Polz Martin F. , Lim Eelin
Published: September 2004

The abundance of
Vibrio vulnificus
in coastal environments has been linked to water temperature, while its relationship to salinity is less clear. We have developed a culture-independent, most-probable-number quantitative PCR approach to examine
V. vulnificus
population dynamics in Barnegat Bay, N.J. Based on the combined analysis of our results from Barnegat Bay and from the literature, the present data show that (i)
V. vulnificus
population dynamics are strongly correlated to water temperature and (ii) although the general trend is for
V. vulnificus
abundance to be inversely correlated with salinity, this relationship depends on salinity levels. Irrespective of temperature, high abundances of
V. vulnificus
are observed at 5 to 10 ppt, which thus appears to be the optimal salinity regime for their survival. At 20 to 25 ppt,
V. vulnificus
abundances show a positive correlation to salinity. Unsuccessful attempts to resuscitate
V. vulnificus
, combined with our inability to detect cells during the winter despite an assay adapted to detect viable but nonculturable (VBNC) cells, suggest that the decline and eventual disappearance of
V. vulnificus
from the water column during the winter months is due primarily to a significant reduction in population size and is not only the consequence of cells entering the VBNC state. These findings are in line with the hypothesis that the sediment serves as a refuge for a subpopulation of
V. vulnificus
over the winter and weather-driven mixing events during the spring initiate a summer bloom in the water column.
Cloning and Characterization of Gluconolactone Oxidase of
Penicillium cyaneo-fulvum
ATCC 10431 and Evaluation of Its Use for Production of
d
-Erythorbic Acid in Recombinant
Pichia pastoris
Authors: Salusjärvi Tuomas , Kalkkinen Nisse , Miasnikov Andrei N.
Published: September 2004

A
d
-erythorbic acid-forming soluble flavoprotein, gluconolactone oxidase (GLO), was purified from
Penicillium cyaneo-fulvum
strain ATCC 10431 and partially sequenced. Peptide sequences were used to isolate a cDNA clone encoding the enzyme. The cloned gene (accession no.
AY576053
) exhibits high levels of similarity with the genes encoding other known eukaryotic lactone oxidases and also with the genes encoding some putative prokaryotic lactone oxidases. Analysis of the coding sequence of the GLO gene indicated the presence of a typical secretion signal sequence at the N terminus of GLO. No other targeting or anchoring signals were found, suggesting that GLO is the first known lactone oxidase that is secreted rather than targeted to the membranes of the endoplasmic reticulum or mitochondria. Experimental evidence, including the N-terminal sequence of mature GLO and data on glycosylation and localization of the enzyme in native and recombinant hosts, supports this analysis. The GLO gene was expressed in
Pichia pastoris
, and recombinant GLO was produced by using the strong methanol-induced
AOX1
promoter. In order to evaluate the suitability of purified GLO for production of
d
-erythorbic acid, we immobilized it on
N
-hydroxysuccinimide-activated Sepharose and found that the immobilized GLO retained full activity during immobilization but was rather unstable under reaction conditions. Our results show that both soluble and immobilized forms of GLO can, in principle, be used for production of
d
-erythorbic acid from
d
-glucono-δ-lactone or (in combination with glucose oxidase and catalase) from glucose. We also demonstrated the feasibility of glucose-
d
-erythorbic acid fermentation with recombinant strains coexpressing GLO and glucose oxidase genes, and we analyzed problems associated with construction of efficient
d
-erythorbic acid-producing hosts.
Characterization of a Highly Enriched
Dehalococcoides
-Containing Culture That Grows on Vinyl Chloride and Trichloroethene
Authors: Duhamel Melanie , Mo Kaiguo , Edwards Elizabeth A.
Published: September 2004

A highly enriched culture that reductively dechlorinates trichloroethene (TCE),
cis
-1,2-dichloroethene (cDCE), and vinyl chloride (VC) to ethene without methanogenesis is described. The
Dehalococcoides
strain in this enrichment culture had a yield of (5.6 ± 1.4) × 10
8
16S rRNA gene copies/μmol of Cl
−
when grown on VC and hydrogen. Unlike the other VC-degrading cultures described in the literature, strains VS and BAV1, this culture maintained the ability to grow on TCE with a yield of (3.6 ± 1.3) × 10
8
16S rRNA gene copies/μmol of Cl
−
. The yields on an electron-equivalent basis measured for the culture grown on TCE and on VC were not significantly different, indicating that both substrates supported growth equally well. PCR followed by denaturing gradient gel electrophoresis, cloning, and phylogenetic analyses revealed that this culture contained one
Dehalococcoides
16S rRNA gene sequence, designated KB-1/VC, that was identical (over 1,386 bp) to the sequences of previously described organisms FL2 and CBDB1. A second
Dehalococcoides
sequence found in separate KB-1 enrichment cultures maintained on cDCE, TCE, and tetrachloroethene was no longer present in the VC-H
2
enrichment culture. This second
Dehalococcoides
sequence was identical to that of BAV1. As neither FL2 nor CBDB1 can dechlorinate VC to ethene in a growth-related fashion, it is clear that current 16S rRNA gene-based analyses do not provide sufficient information to distinguish between metabolically diverse members of the
Dehalococcoides
group.
Electrophoretic Mobility of
Mycobacterium avium
Complex Organisms
Authors: Lytle Darren , Frietch Christy , Covert Terry
Published: September 2004

The electrophoretic mobilities (EPMs) of 30
Mycobacterium avium
complex organisms were measured. The EPMs of 15 clinical isolates ranged from −1.9 to −5.0 μm cm V
−1
s
−1
, and the EPMs of 15 environmental isolates ranged from −1.9 to −4.6 μm cm V
−1
s
−1
at pH 7.
Inhibition of
Listeria monocytogenes
in Fish and Meat Systems by Use of Oregano and Cranberry Phytochemical Synergies
Authors: Lin Y. T. , Labbe R. G. , Shetty Kalidas
Published: September 2004

Optimized phenolics from oregano and cranberry extracts were evaluated for antimicrobial activity against
Listeria monocytogenes
in laboratory media and in beef and fish. The antimicrobial activity increased when oregano and cranberry extracts were mixed at a ratio of 75% oregano and 25% cranberry (wt/wt) with 0.1 mg of phenolic per disk or ml, and the efficacy was further enhanced by lactic acid. The inhibition by phytochemical and lactic acid synergies was most effective when beef and fish slices were stored at 4°C.
Rapid Estimation of Numbers of Fecal
Bacteroidetes
by Use of a Quantitative PCR Assay for 16S rRNA Genes
Authors: Dick Linda K. , Field Katharine G.
Published: September 2004

Assessment of health risk associated with fecal pollution requires a reliable fecal indicator and a rapid quantification method. We report the development of a
Taq
nuclease assay for enumeration of 16S rRNA genes of
Bacteroidetes
. Sensitivity and correlation with standard fecal indicators provide experimental evidence for application of the assay in monitoring fecal pollution.
PCR-Based Identification of Hyperthermophilic Archaea of the Family
Thermococcaceae
Authors: Slobodkina Galina B. , Chernyh Nikolai A. , Slobodkin Alexander I. , Subbotina Irina V. , Bonch-Osmolovskaya Elizaveta A. , Lebedinsky Alexander V.
Published: September 2004

A method for rapid detection and identification of hyperthermophilic archaea of the family
Thermococcaceae
based on PCR amplification of 16S rRNA gene fragments with primers TcPc 173F (5′-TCCCCCATAGGYCTGRGGTACTGGAAGGTC-3′) and TcPc 589R (5′-GCCGTGRGATTTCGCCAGGGACTTACGGGC-3′) was developed and used for identification of new isolates.
Canonical transient receptor potential TRPC7 can function as both a receptor- and store-operated channel in HEK-293 cells
Authors: Lièvremont Jean-Philippe , Bird Gary St. J. , Putney James W.
Published: September 2004

Previous studies on the activation mechanism of canonical transient receptor potential (TRPC) channels have often produced conflicting conclusions. All seven have been shown to be activated by phospholipase C (PLC)-coupled receptors, but TRPC1, TRPC2, TRPC3, TRPC4, TRPC5, and TRPC7 have also been proposed to function as store-operated channels. 1 1 Although PLC activation inevitably leads to activation of store-operated channels, in this report when we refer to PLC-activated channels, we mean those channels that are specifically activated by PLC independently of store depletion. In the case of TRPC3, the expression environment and the expression level appear to determine the mode of regulation. Evidence of a close structural relative of TRPC3, TRPC7, has been presented that this channel is activated by receptor activation or by store depletion. On the basis of previous findings for TRPC3, we reasoned that subtle differences in structure or expression conditions might account for the apparent distinct gating mechanisms of TRPC7. To reexamine the mode of activation of TRPC7, we stably and transiently transfected human embryonic kidney (HEK)-293 cells with cDNA encoding for human TRPC7. We examined the ability of a PLC-activating agonist and an intracellular Ca2+ store-depleting agent to activate these channels. Our findings demonstrate that when transiently expressed in HEK-293 cells, TRPC7 forms channels that are activated by PLC-stimulating agonists, but not by Ca2+ store depletion. However, when stably expressed in HEK-293 cells, TRPC7 can be activated by either Ca2+ store depletion or PLC activation. To our knowledge, this is the first demonstration of a channel protein that can be activated by both receptor- and store-operated modes in the same cell. In addition, the results reconcile the apparently conflicting findings of other laboratories regarding TRPC7 regulation.