bluuebunny/crossref_metadata_2025_split huggingface dataset:
Obligate Sulfide-Dependent Degradation of Methoxylated Aromatic Compounds and Formation of Methanethiol and Dimethyl Sulfide by a Freshwater Sediment Isolate,Parasporobacterium paucivoransgen. nov., sp. nov
Authors: Lomans Bart P. , Leijdekkers Pieter , Wesselink Jan-Jaap , Bakkes Patrick , Pol Arjan , van der Drift Chris , den Camp Huub J. M. Op 
Published: July 2002

ABSTRACTMethanethiol (MT) and dimethyl sulfide (DMS) have been shown to be the dominant volatile organic sulfur compounds in freshwater sediments. Previous research demonstrated that in these habitats MT and DMS are derived mainly from the methylation of sulfide. In order to identify the microorganisms that are responsible for this type of MT and DMS formation, several sulfide-rich freshwater sediments were amended with two potential methyl group-donating compounds, syringate and 3,4,5-trimethoxybenzoate (0.5 mM). The addition of these methoxylated aromatic compounds resulted in excess accumulation of MT and DMS in all sediment slurries even though methanogenic consumption of MT and DMS occurred. From one of the sediment slurries tested, a novel anaerobic bacterium was isolated with syringate as the sole carbon source. The strain, designatedParasporobacterium paucivorans, produced MT and DMS from the methoxy groups of syringate. The hydroxylated aromatic residue (gallate) was converted to acetate and butyrate. LikeSporobacterium olearium, another methoxylated aromatic compound-degrading bacterium, the isolate is a member of the XIVa cluster of the low-GC-contentClostridialesgroup. However, the new isolate differs from all other known methoxylated aromatic compound-degrading bacteria because it was able to degrade syringate in significant amounts only in the presence of sulfide.
Phylogenetic Diversity of Marine Cyanophage Isolates and Natural Virus Communities as Revealed by Sequences of Viral Capsid Assembly Protein Gene g20
Authors: Zhong Yan , Chen Feng , Wilhelm Steven W. , Poorvin Leo , Hodson Robert E. 
Published: July 2002

ABSTRACT

In order to characterize the genetic diversity and phylogenetic affiliations of marine cyanophage isolates and natural cyanophage assemblages, oligonucleotide primers CPS1 and CPS8 were designed to specifically amplify ca. 592-bp fragments of the gene for viral capsid assembly protein g20. Phylogenetic analysis of isolated cyanophages revealed that the marine cyanophages were highly diverse yet more closely related to each other than to enteric coliphage T4. Genetically related marine cyanophage isolates were widely distributed without significant geographic segregation (i.e., no correlation between genetic variation and geographic distance). Cloning and sequencing analysis of six natural virus concentrates from estuarine and oligotrophic offshore environments revealed nine phylogenetic groups in a total of 114 different g20 homologs, with up to six clusters and 29 genotypes encountered in a single sample. The composition and structure of natural cyanophage communities in the estuary and open-ocean samples were different from each other, with unique phylogenetic clusters found for each environment. Changes in clonal diversity were also observed from the surface waters to the deep chlorophyll maximum layer in the open ocean. Only three clusters contained known cyanophage isolates, while the identities of the other six clusters remain unknown. Whether or not these unidentified groups are composed of bacteriophages that infect different
Synechococcus
groups or other closely related cyanobacteria remains to be determined. The high genetic diversity of marine cyanophage assemblages revealed by the g20 sequences suggests that marine viruses can potentially play important roles in regulating microbial genetic diversity.
Purification and Characterization of a Surface Protein from
Lactobacillus fermentum
104R That Binds to Porcine Small Intestinal Mucus and Gastric Mucin
Authors: Rojas Maurilia , Ascencio Felipe , Conway Patricia L. 
Published: July 2002

ABSTRACT

An adhesion-promoting protein involved in the binding of
Lactobacillus fermentum
strain 104R to small intestinal mucus from piglets and to partially purified gastric mucin was isolated and characterized. Spent culture supernatant fluid and bacterial cell wall extracts were fractionated by ammonium sulfate precipitation and gel filtration. The active fraction was purified by affinity chromatography. The adhesion-promoting protein was detected in the fractions by adhesion inhibition and dot blot assays and visualized by polyacrylamide gel electrophoresis (PAGE), sodium dodecyl sulfate-PAGE, and Western blotting with horseradish peroxidase-labeled mucus and mucin. The active fraction was characterized by estimating the relative molecular weight and by assessing the presence of carbohydrates in, and heat sensitivity of, the active region of the adhesion-promoting protein. The purified protein was digested with porcine trypsin, and the peptides were purified in a SMART system. The peptides were tested for adhesion to horseradish peroxidase-labeled mucin by using the dot blot adhesion assay. Peptides which bound mucin were sequenced. It was shown that the purified adhesion-promoting protein on the cell surface of
L. fermentum
104R is extractable with 1 M LiCl and low concentrations of lysozyme but not with 0.2 M glycine. The protein could be released to the culture supernatant fluid after 24 h of growth and had affinity for both small intestinal mucus and gastric mucin. In the native state this protein was variable in size, and it had a molecular mass of 29 kDa when denatured. The denatured protein did not contain carbohydrate moieties and was not heat sensitive. Alignment of amino acids of the adhering peptides with sequences deposited in the EMBL data library showed poor homology with previously published sequences. The protein represents an important molecule for development of probiotics.
Rapid Detection and Enumeration of
Naegleria fowleri
in Surface Waters by Solid-Phase Cytometry
Authors: Pougnard Claire , Catala Philippe , Drocourt Jean-Louis , Legastelois Stephane , Pernin Pierre , Pringuez Emmanuelle , Lebaron Philippe 
Published: July 2002

ABSTRACT

A new method for the rapid and accurate detection of pathogenic
Naegleria fowleri
amoebae in surface environmental water was developed. The method is based on an immunofluorescent assay combined with detection by solid-phase cytometry. In this study we developed and compared two protocols using different reporter systems conjugated to antibodies. The monoclonal antibody Ac5D12 was conjugated with biotin and horseradish peroxidase, and the presence of cells was revealed with streptavidin conjugated to both R-phycoerythrin and cyanine Cy5 (RPE-Cy5) and tyramide-fluorescein isothiocyanate, respectively. The RPE-Cy5 protocol was the most efficient protocol and allowed the detection of both trophozoite and cyst forms in water. The direct counts obtained by this new method were not significantly different from those obtained by the traditional culture approach, and results were provided within 3 h. The sensitivity of the quantitative method is 200 cells per liter. The limit is due only to the filtration capacity of the membrane used.
T-Helper-Cell Proliferative Responses to Whole-Killed Human Immunodeficiency Virus Type 1 (HIV-1) and p24 Antigens of Different Clades in HIV-1-Infected Subjects Vaccinated with HIV-1 Immunogen (Remune)
Authors: Moss Ronald B. , Giermakowska Wieslawa , Wallace Mark R. , Savary Jay , Jensen Fred , Carlo Dennis J. 
Published: July 2002

ABSTRACT
The discovery of multiple subtypes of human immunodeficiency virus type 1 (HIV-1) worldwide has created new challenges for the development of both therapeutic and preventive AIDS vaccines. We examined T-helper proliferative responses to HIV-1 clade A, B, C, G, and E whole-killed virus and to HIV-1 clade G and B core (p24) antigens in HIV-1-infected subjects taking potent antiviral drugs who received HIV immunogen (Remune) therapeutic vaccination. Subjects who were immunized mounted strong proliferative responses to both whole virus and core antigens of the different clades. These results suggest that a whole-killed immunogen may have broad applications as a therapeutic as well as a preventive vaccine in the current multiclade HIV-1 pandemic.
Naive Human T Cells Develop into Th1 Effectors after Stimulation withMycobacterium tuberculosis-Infected Macrophages or Recombinant Ag85 Proteins
Authors: Russo Donna M. , Kozlova Natalia , Lakey David L. , Kernodle Douglas 
Published: July 2002

ABSTRACTMost studies of human T-cell responses in tuberculosis have focused on persons with either active disease or latent infection. Although this work has been critical in defining T-cell correlates of successful versus failed host containment, little is known about the development ofMycobacterium-specific T-cell responses in uninfected persons. To explore this issue, naive T cells from uninfected donors were sensitized in vitro with avirulentMycobacterium tuberculosis-infected autologous macrophages. T-cell lines primed in this manner proliferated and produced cytokines after challenge with mycobacterial antigens. Of 11 such lines, 8 were high Th1 responders, 2 were low Th1 responders, and 1 was a Th2 responder. Furthermore, similar patterns and magnitudes of proliferative and cytokine responses were seen whenMycobacteriuminfection-primed lines were challenged with recombinant antigen 85 (Ag85) proteins. The addition of interleukin 12 (IL-12) during the initial sensitization increased the magnitude of Th1 responses; however, antibody to IL-12 did not eliminate Th1 responses, suggesting that additional factors contributed to the differentiation of these cells. Finally, in the presence of IL-12, recombinant Ag85B was able to prime naive T cells for Th1 responses upon challenge withMycobacterium-infected macrophages or Ag85B. Therefore, under the appropriate conditions, priming with whole bacteria or a subunit antigen can stimulateMycobacterium-specific Th1 effector cell development. Further definition of the antigens and conditions required to drive naive human T cells to differentiate into Th1 effectors should facilitate the development of an improved tuberculosis vaccine.
Human Monocyte Receptors Involved in Tumor Necrosis Factor Responses to Group B Streptococcal Products
Authors: Cuzzola Maria , Mancuso Giuseppe , Beninati Concetta , Biondo Carmelo , von Hunolstein Christina , Orefici Graziella , Espevik Terje , Flo Trude H. , Teti Giuseppe 
Published: July 2002

ABSTRACTSeveral group B streptococcal products have been previously found to stimulate human monocytes to produce tumor necrosis factor alpha. In order to identify the receptors involved in these responses, monocytes were stimulated with purified group- or type-specific carbohydrates or lipoteichoic acid in the presence of anti-receptor monoclonal antibodies, soluble CD14, or lipopolysaccharide-binding protein. Results indicate that CD14 plays an important role in tumor necrosis factor alpha responses to all of the stimuli tested. Moreover, both CD14 and complement receptor type 3 may be involved in responses to the group-antigen.
Innate Lung Defenses and CompromisedPseudomonas aeruginosaClearance in the Malnourished Mouse Model of Respiratory Infections in Cystic Fibrosis
Authors: Yu H. , Nasr S. Z. , Deretic V. 
Published: July 2002

ABSTRACTCystic fibrosis (CF) is characterized by dysfunction of the digestive and respiratory tracts resulting in generalized malnutrition and chronic respiratory infections. Chronic lung infections withPseudomonas aeruginosa, intense neutrophil-dominated airway inflammation, and progressive lung disease are the major cause of high morbidity and mortality in CF. Here we investigated the effects of malnutrition in CF on innate lung defenses, susceptibility toP. aeruginosacolonization, and associated inflammation, using aerosol models of acute and chronic infections in normal, malnourished, and transgenic mice.CFTRm1Unc−/−knockout mice displayed body weight variations and showed variable pulmonary clearance ofP. aeruginosa. This variability was not detected in bitransgenicCFTRm1Unc−/−(FABP-hCFTR) mice in which the intestinal defect had been corrected. Diet-induced protein calorie malnutrition in C57BL/6J mice resulted in impaired pulmonary clearance ofP. aeruginosa. Tumor necrosis factor alpha (TNF-α) and nitrite levels detected upon exposure toP. aeruginosaaerosols were lower in the lungs of the malnourished C57BL/6J mice relative than in lungs of mice fed a normal diet. The role of TNF-α and reactive nitrogen intermediates inP. aeruginosaclearance was tested in TNF-α and inducible nitric oxide synthase (iNOS) knockout mice.P. aeruginosaclearance was diminished in transgenic TNF-α- and iNOS-deficient mice. In contrast to the effects of TNF-α and iNOS, gamma interferon knockout mice retained a full capacity to eliminateP. aeruginosafrom the lung. Malnutrition also contributed to excessive inflammation in C57BL/6J mice upon chronic challenge withP. aeruginosa. The repeatedly infected malnourished host did not produce interleukin-10, a major anti-inflammatory cytokine absent or diminished in the bronchoalveolar fluids of CF patients. These results are consistent with a model in which defectiveCFTRin the intestinal tract leads to nutritional deficiency which in turn contributes to compromised innate lung defenses, bacterial colonization, and excessive inflammation in the CF respiratory tract.
Transforming Growth Factor-β Inhibits Lipopolysaccharide-Stimulated Expression of Inflammatory Cytokines in Mouse Macrophages through Downregulation of Activation Protein 1 and CD14 Receptor Expression
Authors: Imai Kenichi , Takeshita Akira , Hanazawa Shigemasa 
Published: July 2002

ABSTRACTThe septic shock that occurs in gram-negative infections is caused by a cascade of inflammatory cytokines. Several studies showed that transforming growth factor-β1 (TGF-β1) inhibits this septic shock through suppression of expression of the lipopolysaccharide (LPS)-induced inflammatory cytokines. In this study, we investigated whether TGF-β1 inhibition of LPS-induced expression of inflammatory cytokines in the septic shock results from downregulation of LPS-stimulated expression of CD14, an LPS receptor. TGF-β1 markedly inhibited LPS stimulation of CD14 mRNA and protein levels in mouse macrophages. LPS-stimulated expression of CD14 was dramatically inhibited by addition of antisense, but not sense, c-fosand c-junoligonucleotides. Since TGF-β1 pretreatment inhibited LPS-stimulated expression of c-fosand c-jungenes and also the binding of nuclear proteins to the consensus sequence of the binding site for activation protein 1 (AP-1), a heterodimer of c-Fos and c-Jun, in the cells, TGF-β1 inhibition of CD14 expression may be a consequence of downregulation of AP-1. LPS-stimulated expression of interleukin-1β and tumor necrosis factor alpha genes in the cells was inhibited by addition of CD14 antisense oligonucleotide. Also, TGF-β1 inhibited the LPS-stimulated production of both inflammatory cytokines by the macrophages. In addition, TGF-β1 inhibited expression of the two cytokines in several organs of mice receiving LPS. Thus, our results suggest that TGF-β1 inhibition of LPS-stimulated inflammatory responses resulted from downregulation of CD14 and also may be a possible mechanism of TGF-β1 inhibition of LPS-induced septic shock.
Molecular Cloning and Expression of Cu/Zn-Containing Superoxide Dismutase from
Fasciola hepatica
Authors: Kim Tong-Soo , Jung Younghun , Na Byoung-Kuk , Kim Ki-Sun , Chung Pyung-Rim 
Published: July 2002

ABSTRACT

The cytosolic superoxide dismutase (SOD) of
Fasciola hepatica
, a causative agent of fascioliasis, was purified and characterized. The enzyme consists of two identical subunits, each with an apparent molecular mass of 17.5 kDa. An analysis of the enzyme's primary structure and inhibition studies revealed that the enzyme is a copper/zinc-containing SOD (Cu/Zn-SOD). The enzyme activity was relatively stable in a broad pH range, from pH 7.0 to 10.0, and the enzyme showed maximum activity at pH 7.5. This enzyme also displayed strong antigenicity against sera of bovine and human subjects with fascioliasis. The SOD gene fragment was amplified by PCR with degenerate oligonucleotide primers derived from amino acid sequences conserved in the Cu/Zn-SODs of other organisms. An
F. hepatica
cDNA library was screened with the SOD gene fragment as a probe. As a result, a complete gene encoding the Cu/Zn-SOD was identified, and its nucleotide sequence was determined. The gene had an open reading frame of 438 bp and 146 deduced amino acids. Comparison of the deduced amino acid sequence of the enzyme with previously reported Cu/Zn-SOD amino acid sequences revealed considerably high homologies. The coding region of the
F. hepatica
Cu/Zn-SOD was cloned and expressed in
Escherichia coli
. Staining of native polyacrylamide gel for SOD activity of the expressed protein revealed SOD activity that was inactivated by potassium cyanide and hydrogen peroxide but not by sodium azide. This means that the presence of the recombinant fusion protein is indicative of Cu/Zn-SOD. The expressed protein also reacted with sera of bovine and human subjects with fascioliasis, but it did not react with sera of uninfected bovine and human subjects.
Mutator Natural
Escherichia coli
Isolates Have an Unusual Virulence Phenotype
Authors: Picard Bertrand , Duriez Patrick , Gouriou Stéphanie , Matic Ivan , Denamur Erick , Taddei François 
Published: July 2002

ABSTRACT

A small percentage of natural
Escherichia coli
isolates (both commensal and pathogenic) have a mutator phenotype related to defects in methyl-directed mismatch repair (MR) genes. We investigated whether there was a direct link between the mutator phenotype and virulence by (i) studying the relationships between mutation rate and virulence in a mouse model of extraintestinal virulence for 88 commensal and extraintestinal pathogenic
E. coli
isolates and (ii) comparing the virulence in mice of MR-deficient and MR-proficient strains that were otherwise isogenic. The results provide no support for the hypothesis that the mutator phenotype has a direct role in virulence or is associated with increased virulence. Most of the natural mutator strains studied displayed an unusual virulence phenotype with (i) a lack of correspondence between the number of virulence determinants and pathogenicity in mice and (ii) an intermediate level of virulence. On a large evolutionary scale, the mutator phenotype may help parasites to achieve an intermediate rate of virulence which mathematical models predict to be selected for during long-term parasite-host interactions.
Outer Membrane Targeting of Passenger Proteins by the Vacuolating Cytotoxin Autotransporter of
Helicobacter pylori
Authors: Fischer Wolfgang , Buhrdorf Renate , Gerland Elke , Haas Rainer 
Published: July 2002

ABSTRACT

Helicobacter pylori
produces a number of proteins associated with the outer membrane, including adhesins and the vacuolating cytotoxin. These proteins are supposed to integrate into the outer membrane by β-barrel structures, characteristic of the family of autotransporter proteins. By using the SOMPES (shuttle vector-based outer membrane protein expression) system for outer membrane protein production, we were able to functionally express in
H. pylori
the cholera toxin B subunit genetically fused to the C-terminal VacA domain. We demonstrate that the fusion protein is translocated to the
H. pylori
outer membrane and that the CtxB domain is exposed on the
H. pylori
surface. Thus, we provide the first experimental evidence that the C-terminal β-domain of VacA can transport a foreign passenger protein to the
H. pylori
surface and hence acts as a functional autotransporter.
Comparison of Genetic Divergence and Fitness between Two Subclones of
Helicobacter pylori
Authors: Björkholm Britta , Lundin Annelie , Sillén Anna , Guillemin Karen , Salama Nina , Rubio Carlos , Gordon Jeffrey I. , Falk Per , Engstrand Lars 
Published: July 2002

ABSTRACT

Helicobacter pylori
has a very plastic genome, reflecting its high rate of recombination and point mutation. This plasticity promotes divergence of the population by the development of subclones and presumably enhances adaptation to host niches. We have investigated the genotypic and phenotypic characteristics of two such subclones isolated from one patient as well as the genetic evolution of these isolates during experimental infection. Whole-genome genotyping of the isolates using DNA microarrays revealed that they were more similar to each other than to a panel of other genotyped strains recovered from different hosts. Nonetheless, they still showed significant differences. For example, one isolate (67:21) contained the entire Cag pathogenicity island (PAI), whereas the other (67:20) had excised the PAI. Phenotypic studies disclosed that both isolates expressed adhesins that recognized human histo-blood group Lewis
b
glycan receptors produced by gastric pit and surface mucus cells. In addition, both isolates were able to colonize, to equivalent density and with similar efficiency, germ-free transgenic mice genetically engineered to synthesize Lewis
b
glycans in their pit cells (12 to 14 mice/isolate). Remarkably, the Cag PAI-negative isolate was unable to colonize conventionally raised Lewis
b
transgenic mice harboring a normal gastric microflora, whereas the Cag PAI-positive isolate colonized 74% of the animals (39 to 40 mice/isolate). The genomic evolution of both isolates during the infection of conventionally raised and germ-free mice was monitored over the course of 3 months. The Cag PAI-positive isolate was also surveyed after a 10 month colonization of conventionally raised transgenic animals (
n
= 9 mice). Microarray analysis of the Cag PAI and sequence analysis of the
cagA
,
recA
, and 16S rRNA genes disclosed no changes in recovered isolates. Together, these results reveal that the
H. pylori
population infecting one individual can undergo significant divergence, creating stable subclones with substantial genotypic and phenotypic differences.
Mucosal Vaccination with Recombinantly Attenuated Staphylococcal Enterotoxin B and Protection in a Murine Model
Authors: Stiles Bradley G. , Garza Anthony R. , Ulrich Robert G. , Boles James W. 
Published: July 2002

ABSTRACTPrevious work in our laboratory revealed that mice parenterally vaccinated with recombinantly attenuated staphylococcal enterotoxin (SE) or toxic shock syndrome toxin 1 develop protective antibodies against a lethal intraperitoneal (i.p.) toxin challenge. This study investigated the efficacy of nasal and oral immunizations with an SEB vaccine (SEBv) toward an i.p. or mucosal (via an aerosol) toxin challenge. Both vaccination routes, with the immunoadjuvant cholera toxin (CT), elicited comparable SEB-specific immunoglobulin A (IgA) and IgG levels in saliva. Nasal or oral inoculations also generated SEB-specific IgA, IgG, and IgM in the serum, but the nasal route yielded higher specific IgG titers. SEBv alone, when given nasally or orally, did not induce any detectable SEB-specific antibody. Mice vaccinated mucosally were protected against a 50% lethal dose of wild-type SEB given i.p. or mucosally, thus demonstrating that nasal or oral administration of this SEBv, with CT, elicits systemic and mucosal antibodies to SEB that protect against SEB-induced lethal shock.