bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of Genetically Modified
Pseudomonas putida
WCS358r on the Fungal Rhizosphere Microflora of Field-Grown Wheat
Authors: Glandorf Debora C. M. , Verheggen Patrick , Jansen Timo , Jorritsma Jan-Willem , Smit Eric , Leeflang Paula , Wernars Karel , Thomashow Linda S. , Laureijs Eric , Thomas-Oates Jane E. , Bakker Peter A. H. M. , van Loon Leendert C. 
Published: July 2002

ABSTRACT

We released genetically modified
Pseudomonas putida
WCS358r into the rhizospheres of wheat plants. The two genetically modified derivatives, genetically modified microorganism (GMM) 2 and GMM 8, carried the
phz
biosynthetic gene locus of strain
P. fluorescens
2-79 and constitutively produced the antifungal compound phenazine-1-carboxylic acid (PCA). In the springs of 1997 and 1998 we sowed wheat seeds treated with either GMM 2, GMM 8, or WCS358r (approximately 10
7
CFU per seed), and measured the numbers, composition, and activities of the rhizosphere microbial populations. During both growing seasons, all three bacterial strains decreased from 10
7
CFU per g of rhizosphere sample to below the limit of detection (10
2
CFU per g) 1 month after harvest of the wheat plants. The
phz
genes were stably maintained, and PCA was detected in rhizosphere extracts of GMM-treated plants. In 1997, but not in 1998, fungal numbers in the rhizosphere, quantified on 2% malt extract agar (total filamentous fungi) and on Komada's medium (mainly
Fusarium
spp.), were transiently suppressed in GMM 8-treated plants. We also analyzed the effects of the GMMs on the rhizosphere fungi by using amplified ribosomal DNA restriction analysis. Introduction of any of the three bacterial strains transiently changed the composition of the rhizosphere fungal microflora. However, in both 1997 and 1998, GMM-induced effects were distinct from those of WCS358r and lasted for 40 days in 1997 and for 89 days after sowing in 1998, whereas effects induced by WCS358r were detectable for 12 (1997) or 40 (1998) days. None of the strains affected the metabolic activity of the soil microbial population (substrate-induced respiration), soil nitrification potential, cellulose decomposition, plant height, or plant yield. The results indicate that application of GMMs engineered to have improved antifungal activity can exert nontarget effects on the natural fungal microflora.
Impact of 2,4-Diacetylphloroglucinol-Producing Biocontrol Strain
Pseudomonas fluorescens
F113 on Intraspecific Diversity of Resident Culturable Fluorescent Pseudomonads Associated with the Roots of Field-Grown Sugar Beet Seedlings
Authors: Moënne-Loccoz Yvan , Tichy Hans-Volker , O'Donnell Anne , Simon Reinhard , O'Gara Fergal 
Published: July 2002

ABSTRACT

The impact of the 2,4-diacetylphloroglucinol-producing biocontrol agent
Pseudomonas fluorescens
F113Rif on the diversity of the resident community of culturable fluorescent pseudomonads associated with the roots of field-grown sugar beet seedlings was evaluated. At 19 days after sowing, the seed inoculant F113Rif had replaced some of the resident culturable fluorescent pseudomonads at the rhizoplane but had no effect on the number of these bacteria in the rhizosphere. A total of 498 isolates of resident fluorescent pseudomonads were obtained and characterized by molecular means at the level of broad phylogenetic groups (by amplified ribosomal DNA restriction analysis) and at the strain level (with random amplified polymorphic DNA markers) as well as phenotypically (55 physiological tests). The introduced pseudomonad induced a major shift in the composition of the resident culturable fluorescent
Pseudomonas
community, as the percentage of rhizoplane isolates capable of growing on three carbon substrates (erythritol, adonitol, and
l
-tryptophan) not assimilated by the inoculant was increased from less than 10% to more than 40%. However, the pseudomonads selected did not display enhanced resistance to 2,4-diacetylphloroglucinol. The shift in the resident populations, which was spatially limited to the surface of the root (i.e., the rhizoplane), took place without affecting the relative proportions of phylogenetic groups or the high level of strain diversity of the resident culturable fluorescent
Pseudomonas
community. These results suggest that the root-associated
Pseudomonas
community of sugar beet seedlings is resilient to the perturbation that may be caused by a taxonomically related inoculant.
Putative Virulence Factor Expression by Clinical and Food Isolates of
Bacillus
spp. after Growth in Reconstituted Infant Milk Formulae
Authors: Rowan Neil J. , Deans Karen , Anderson John G. , Gemmell Curtis G. , Hunter Iain S. , Chaithong Thararat 
Published: July 2002

ABSTRACT

Forty-seven strains representing 14 different
Bacillus
species isolated from clinical and food samples were grown in reconstituted infant milk formulae (IMF) and subsequently assessed for adherence to, invasion of, and cytotoxicity toward HEp-2 and Caco-2 cells. Cell-free supernatant fluids from 38 strains (81%) were shown to be cytotoxic, 43 strains (91%) adhered to the test cell lines, and 23 strains (49%) demonstrated various levels of invasion. Of the 21
Bacillus cereus
strains examined, 5 (24%) were invasive. A larger percentage of clinically derived
Bacillus
species (20%) than of similar species tested from the food environment were invasive. Increased invasion occurred after growth of selected
Bacillus
species in reconstituted IMF containing glucose. While PCR primer studies revealed that many different
Bacillus
species contained DNA sequences encoding the hemolysin BL (HBL) enterotoxin complex and
B. cereus
enterotoxin T, not all of these isolates expressed these diarrheagenic genes after growth in reconstituted IMF. Of the 47
Bacillus
isolates examined, 3 isolates of
B. cereus
and 1 isolate of
B. subtilis
produced the HBL enterotoxin after 18 h of growth in brain heart infusion broth. However, eight isolates belonging to the species
B. cereus
,
B. licheniformis
,
B. circulans
, and
B. megaterium
were found to produce this enterotoxin after growth in reconstituted IMF when assessed with the
B. cereus
enterotoxin (diarrheal type) reversed passive latex agglutination (RPLA) kit. It is concluded that several
Bacillus
species occurring occasionally in clinical specimens and food samples are of potential medical significance due to the expression of putative virulence factors.
Rapid and Efficient Extraction Method for Reverse Transcription-PCR Detection of Hepatitis A and Norwalk-Like Viruses in Shellfish
Authors: Kingsley David H. , Richards Gary P. 
Published: July 2002

ABSTRACT

As part of an effort to develop a broadly applicable test for Norwalk-like viruses and hepatitis A virus (HAV) in shellfish, a rapid extraction method that is suitable for use with one-step reverse transcription (RT)-PCR-based detection methods was developed. The method involves virus extraction using a pH 9.5 glycine buffer, polyethylene glycol (PEG) precipitation, Tri-reagent, and purification of viral poly(A) RNA by using magnetic poly(dT) beads. This glycine–PEG–Tri-reagent–poly(dT) method can be performed in less than 8 h on hard-shell clams (
Mercenaria mercenaria
) and Eastern oysters (
Crassostrea virginica
) and, when coupled with RT-PCR-based detection, can yield results within 24 h. Observed sensitivities for seeded shellfish extracts are as low as 0.015 PFU of HAV and 22.4 RT-PCR
50
units for Norwalk virus. Detection of HAV in live oysters experimentally exposed to contaminated seawater is also demonstrated. An adaptation of this method was used to identify HAV in imported clams (tentatively identified as
Ruditapes philippinarum
) implicated in an outbreak of food-borne viral illness. All of the required reagents are commercially available. This method should facilitate the implementation of RT-PCR testing of commercial shellfish.
Biosorption of Copper by a Bacterial Biofilm on a Flexible Polyvinyl Chloride Conduit
Authors: Qureshi Fouad M. , Badar Uzma , Ahmed Nuzhat 
Published: July 2002

ABSTRACT
Inexpensive technologies with less-than-optimal efficiencies as a strategy for countering economic restraints to pollution control have been evaluated by using a laboratory-scale biotreatment process for copper-containing effluent. Economizing measures include the use of polyvinyl chloride (PVC) cylinders fashioned from commercially available flexible PVC conduit to support a biofilm that was cultured in an inexpensive medium prepared in wastewater. The biofilm was challenged by aqueous copper solution in a bioreactor and subsequently analyzed under a scanning electron microscope with energy-dispersive X-ray microanalysis.
Isolation and Initial Characterization of a Bacterial Consortium Able To Mineralize Fluorobenzene
Authors: Carvalho M.F. , Alves C.C.T. , Ferreira M.I.M. , De Marco P. , Castro P.M.L. 
Published: July 2002

ABSTRACT

Fluorinated compounds are known to be more resistant to microbial degradation than other halogenated chemicals. A microbial consortium capable of aerobic biodegradation of fluorobenzene (FB) as the sole source of carbon and energy was isolated by selective enrichment from sediments collected in a drain near an industrial site. A combination of three microbial strains recovered from the enriched consortium was shown to be necessary for complete FB mineralization. Two of the strains (F1 and F3) were classified by 16S rRNA analysis as belonging to the
Sphingobacterium/Flavobacterium
group, while the third (F4) falls in the β-
Proteobacteria
group, clustering with
Alcaligenes
species. Strain F4 was consistently found in the liquid cultures in a much greater proportion than strains F1 and F3 (86:8:6 for F4, F1, and F3, respectively). Stoichiometric release of fluoride ions was measured in batch and fed-batch cultures. In batch cultures, the consortium was able to use FB up to concentrations of 400 mg liter
−1
and was able to utilize a range of other organic compounds, including 4-fluorophenol and 4-fluorobenzoate. To our knowledge this is the first time biodegradation of FB as a sole carbon source has been reported.
Natural Genetic Transformation of Clinical Isolates of
Escherichia coli
in Urine and Water
Authors: Woegerbauer Markus , Jenni Bernard , Thalhammer Florian , Graninger Wolfgang , Burgmann Heinz 
Published: July 2002

ABSTRACT

Transfer of plasmid-borne antibiotic resistance genes in
Escherichia coli
wild-type strains is possible by transformation under naturally occurring conditions in oligotrophic, aquatic environments containing physiologic concentrations of calcium. In contrast, transformation is suppressed in nitrogen-rich body fluids like urine, a common habitat of uropathogenic strains. Current knowledge indicates that transformation of these
E. coli
wild-type strains is of no relevance for the acquisition of resistance in this clinically important environment.
Sequence Analysis of Marine Virus Communities Reveals that Groups of Related Algal Viruses Are Widely Distributed in Nature
Authors: Short Steven M. , Suttle Curtis A. 
Published: July 2002

ABSTRACT

Algal-virus-specific PCR primers were used to amplify DNA polymerase (
pol
) gene fragments from geographically isolated natural virus communities. Natural algal virus communities were obtained from coastal sites in the Pacific Ocean in British Columbia, Canada, and the Southern Ocean near the Antarctic peninsula. Genetic fingerprints of algal virus communities were generated using denaturing gradient gel electrophoresis (DGGE). Sequencing efforts recovered 33 sequences from the gradient gel. Of the 33 sequences examined, 25 encoded a conserved amino acid motif indicating that the sequences were
pol
gene fragments. Furthermore, the 25
pol
sequences were related to
pol
gene fragments from known algal viruses. In addition, similar virus sequences (>98% sequence identity) were recovered from British Columbia and Antarctica. Results from this study demonstrate that DGGE with degenerate primers can be used to qualitatively fingerprint and assess genetic diversity in specific subsets of natural virus communities and that closely related viruses occur in distant geographic locations. DGGE is a powerful tool for genetically fingerprinting natural virus communities and may be used to examine how specific components of virus communities respond to experimental manipulations.
Rapid Assessment of the Physiological Status of the Polychlorinated Biphenyl Degrader
Comamonas testosteroni
TK102 by Flow Cytometry
Authors: Hiraoka Yoshinori , Kimbara Kazuhide 
Published: July 2002

ABSTRACT

The viability of the polychlorinated biphenyl-degrading bacterium
Comamonas testosteroni
TK102 was assessed by flow cytometry (FCM) with the fluorogenic ester Calcein-AM (CAM) and the nucleic acid dye propidium iodide (PI). CAM stained live cells, whereas PI stained dead cells. When double staining with CAM and PI was performed, three physiological states, i.e., live (calcein positive, PI negative), dead (calcein negative, PI positive), and permeabilized (calcein positive, PI positive), were detected. To evaluate the reliability of this double-staining method, suspensions of live and dead cells were mixed in various proportions and analyzed by FCM. The proportion of dead cells measured by FCM directly correlated with the proportion of dead cells in the sample (
y
= 0.9872
x
+ 0.18;
R
2
= 0.9971). In addition, the proportion of live cells measured by FCM inversely correlated with the proportion of dead cells in the sample (
y
= −0.9776
x
+ 98.36;
R
2
= 0.9962). The proportion of permeabilized cells was consistently less than 2%. These results indicate that FCM in combination with CAM and PI staining is rapid (≤1 h) and distinguishes correctly among live, dead, and permeabilized cells.
Differential Regulation of
mnp2
, a New Manganese Peroxidase-Encoding Gene from the Ligninolytic Fungus
Trametes versicolor
PRL 572
Authors: Johansson Tomas , Nyman Per Olof , Cullen Daniel 
Published: July 2002

ABSTRACT

A peroxidase-encoding gene,
mnp2
, and its corresponding cDNA were characterized from the white-rot basidiomycete
Trametes versicolor
PRL 572. We used quantitative reverse transcriptase-mediated PCR to identify
mnp2
transcripts in nutrient-limited stationary cultures. Although
mnp2
lacks upstream metal response elements (MREs), addition of MnSO
4
to cultures increased
mnp2
transcript levels 250-fold. In contrast, transcript levels of an MRE-containing gene of
T. versicolor
,
mnp1
, increased only eightfold under the same conditions. Thus, the manganese peroxidase genes in
T. versicolor
are differentially regulated, and upstream MREs are not necessarily involved. Our results support the hypothesis that fungal and plant peroxidases arose through an ancient duplication and folding of two structural domains, since we found the
mnp1
and
mnp2
polypeptides to have internal homology.
Fate of
Escherichia coli
O157:H7 in Manure-Amended Soil
Authors: Jiang Xiuping , Morgan Jennie , Doyle Michael P. 
Published: July 2002

ABSTRACT

Escherichia coli
O157:H7 cells survived for up to 77, >226, and 231 days in manure-amended autoclaved soil held at 5, 15, and 21°C, respectively. Pathogen populations declined more rapidly in manure-amended unautoclaved soil under the same conditions, likely due to antagonistic interactions with indigenous soil microorganisms.
E. coli
O157:H7 cells were inactivated more rapidly in both autoclaved and unautoclaved soils amended with manure at a ratio of 1 part manure to 10 parts soil at 15 and 21°C than in soil samples containing dilute amounts of manure. The manure-to-soil ratio, soil temperature, and indigenous microorganisms of the soil appear to be contributory factors to the pathogen's survival in manure-amended soil.
Cloning and Characterization of the
Zymobacter palmae
Pyruvate Decarboxylase Gene (
pdc
) and Comparison to Bacterial Homologues
Authors: Raj Krishnan Chandra , Talarico Lee A. , Ingram Lonnie O. , Maupin-Furlow Julie A. 
Published: July 2002

ABSTRACT

Pyruvate decarboxylase (PDC) is the key enzyme in all homo-ethanol fermentations. Although widely distributed among plants, yeasts, and fungi, PDC is absent in animals and rare in bacteria (established for only three organisms). Genes encoding the three known bacterial
pdc
genes have been previously described and expressed as active recombinant proteins. The
pdc
gene from
Zymomonas mobilis
has been used to engineer ethanol-producing biocatalysts for use in industry. In this paper, we describe a new bacterial
pdc
gene from
Zymobacter palmae
. The pattern of codon usage for this gene appears quite similar to that for
Escherichia coli
genes. In
E. coli
recombinants, the
Z. palmae
PDC represented approximately 1/3 of the soluble protein. Biochemical and kinetic properties of the
Z. palmae
enzyme were compared to purified PDCs from three other bacteria. Of the four bacterial PDCs, the
Z. palmae
enzyme exhibited the highest specific activity (130 U mg of protein
−1
) and the lowest

K
m

for pyruvate (0.24 mM). Differences in biochemical properties, thermal stability, and codon usage may offer unique advantages for the development of new biocatalysts for fuel ethanol production.
Anticardiolipin Antibodies in Patients with Chronic Hepatitis C Virus Infection: Characterization in Relation to Antiphospholipid Syndrome
Authors: Ordi-Ros Josep , Villarreal Julieta , Monegal Francesc , Sauleda Silvia , Esteban Ignacio , Vilardell Miquel 
Published: July 2002

ABSTRACT

The antiphospholipid syndrome (APS) is usually defined by the association of clinical manifestations that comprise venous and/or arterial thrombosis, recurrent fetal losses, and thrombocytopenia, along with the presence of anticardiolipin (aCL) antibodies and/or lupus anticoagulant. Various infectious diseases can induce aCL; however, these antibodies are not usually associated with thrombotic events, as happens with autoimmune diseases, in which these antibodies need the presence of β
2
-glycoprotein I. Levels of immunoglobulin G (IgG) and IgM aCL antibodies were determined by enzyme-linked immunosorbent assay for 243 patients with chronic hepatitis C virus (HCV) infection and 100 healthy controls. Clinical events of APS, the level of β
2
-glycoprotein dependence of aCL, the presence of cryoglobulins and other autoantibodies, and cross-reactivity between purified aCL and HCV were evaluated. Positive results for aCL antibodies were found more frequently (3.3%) for the patients with HCV infection than for healthy controls (0%). All positive aCL antibodies were β
2
-glycoprotein I independent. No significant association was found between aCL antibodies and clinical manifestations of APS, neither was one found between the presence of other autoantibodies or cryoglobulins and that of aCL. Finally, no cross-reactivity between aCL antibodies and HCV antigens was observed. As previously reported, aCL antibodies seem to be an epiphenomenon, and they do not have clinical or laboratory significance in HCV patients.
A Double-Blind, Adjuvant-Controlled Trial of Human Immunodeficiency Virus Type 1 (HIV-1) Immunogen (Remune) Monotherapy in Asymptomatic, HIV-1-Infected Thai Subjects with CD4-Cell Counts of >300
Authors: Churdboonchart V. , Sakondhavat C. , Kulpradist S. , Na Ayudthya B. Isarangkura , Chandeying V. , Rugpao S. , Boonshuyar C. , Sukeepaisarncharoen W. , Sirawaraporn W. , Carlo D. J. , Moss R. 
Published: July 2002

ABSTRACT

We examined the effect of a human immunodeficiency virus (HIV)-specific immune-based therapy in Thailand, where access to antiviral drug therapy is limited. A 40-week trial was conducted with 297 asymptomatic, HIV-infected Thai subjects with CD4-cell counts greater than 300 μl/mm
3
. Subjects were randomized to receive either HIV type 1 (HIV-1) immunogen (Remune; inactivated HIV-1 from which gp120 is depleted in incomplete Freund's adjuvant or adjuvant control at 0, 12, 24, and 36 weeks at five different clinical sites in Thailand. Neither group received antiviral drug therapy. The a priori primary endpoint for the trial was changes in CD4-cell counts with secondary parameters of percent changes in CD8-cell counts (percent CD4, CD8, and CD4/CD8) and body weight. Subsets of subjects were also examined for changes in plasma HIV-1 RNA levels, Western blot immunoreactivity, and HIV-1 delayed-type hypersensitivity (DTH) skin test reactivity. There was a significant difference in changes in CD4-cell counts that favored the HIV-1 immunogen-treated group compared to those for the adjuvant-treated control group (
P
< 0.05). On average, for HIV-1 immunogen-treated subjects CD4-cell counts increased by 84 cells by week 40, whereas the increase for the control group was 38 cells by week 40. This increase in CD4-cell count was associated with increased HIV-specific immunogenicity, as shown by Western blotting and enhanced HIV-1 DTH skin reactivity. No significant differences in adverse events were observed between the groups. The results of this trial suggest that HIV-1 immunogen is safe and significantly increases CD4-cell counts and HIV-specific immunity compared to those achieved with the adjuvant control in asymptomatic HIV-1-infected subjects not taking antiviral drugs.
Fecal Excretion of a Novel Human Circovirus, TT Virus, in Healthy Children
Authors: Lin C.-L. , Kyono W. , Tongson J. , Chua P. K. , Easa D. , Yanagihara R. , Nerurkar V. R. 
Published: July 2002

ABSTRACT
The role of TT virus (TTV) as a human pathogen is unclear, as is the mode of TTV transmission. To determine the prevalence of TTV infection and the possible fecal-oral route of transmission, we analyzed fecal specimens from 67 healthy, nontransfused children for TTV DNA sequences by heminested PCR, using the NG and T primer sets. The overall prevalence of TTV fecal excretion was 22.4% (15 of 67), with the T primer set (19.4%) being more sensitive than the NG primer set (10.4%). TTV prevalence based on gender or ethnicity showed no significant differences. None of seven children in the 0- to 6-month age group had detectable TTV in feces. Of three sets of siblings, two unrelated sets of twins, ages 33 and 37 months, were negative for fecal TTV DNA, while the third set of siblings, ages 99 and 35 months, was positive. The absence of TTV in the feces of children younger than 6 months and the high prevalence (40%) in children 7 to 12 months of age is consistent with age-specific acquisition of TTV infection by the nonparenteral route. TTV genotypes 1, 3, 4, and 5 were represented in our study population. TTV-positive siblings had TTV genotypes 1 and 4, suggesting unrelated environmental sources of TTV infection. This observation suggests a possible time frame for TTV acquisition in children which coincides with increased interaction with their environment and increased susceptibility to infectious agents.
Determination of
Helicobacter pylori
Virulence by Simple Gene Analysis of the
cag
Pathogenicity Island
Authors: Ikenoue Tsuneo , Maeda Shin , Ogura Keiji , Akanuma Masao , Mitsuno Yuzo , Imai Yasuo , Yoshida Haruhiko , Shiratori Yasushi , Omata Masao 
Published: July 2002

ABSTRACT

Nucleic acid amplification was performed for five loci in the
cag
pathogenicity island (PAI) of
Helicobacter pylori
(comprising
cagA
, the
cagA
promoter region,
cagE
,
cagT
, and the left end of
cag
II [LEC]), and gastric inflammation in patients was evaluated. Of 204
H. pylori
isolates from Japanese patients (53 with peptic ulcer, 55 with gastric cancer, and 96 with chronic gastritis), 197 (96.6%) were positive for all five loci. Two isolates (1%) were negative for all five loci, and five isolates (2.4%) were positive for only
cagA
and LEC. These latter seven isolates were all from patients with mild chronic gastritis. Neutrophil infiltration in gastric mucosa was significantly milder in patients infected with partially or totally deleted-PAI strains than in those with intact-PAI strains. The
cagE
gene was a more accurate marker of an intact
cag
PAI than the
cagA
gene, and
cagE
seemed to be more useful in discriminating between
H. pylori
strains causing different rates of disease progression.