bluuebunny/crossref_metadata_2025_split huggingface dataset:
Construction and Analysis of Photolyase Mutants of
Pseudomonas aeruginosa
and
Pseudomonas syringae
: Contribution of Photoreactivation, Nucleotide Excision Repair, and Mutagenic DNA Repair to Cell Survival and Mutability following Exposure to UV-B Radiation
Authors: Kim Jae J. , Sundin George W. 
Published: July 2002

ABSTRACT

Based on nucleotide sequence homology with the
Escherichia coli
photolyase gene (
phr
), the
phr
sequence of
Pseudomonas aeruginosa
PAO1 was identified from the genome sequence, amplified by PCR, cloned, and shown to complement a known
phr
mutation following expression in
Escherichia coli
SY2. Stable, insertional
phr
mutants containing a tetracycline resistance gene cassette were constructed in
P. aeruginosa
PAO1 and
P. syringae
pv. syringae FF5 by homologous recombination and sucrose-mediated counterselection. These mutants showed a decrease in survival compared to the wild type of as much as 19-fold after irradiation at UV-B doses of 1,000 to 1,550 J m
−2
followed by a recovery period under photoreactivating conditions. A
phr uvrA
mutant of
P. aeruginosa
PAO1 was markedly sensitive to UV-B irradiation exhibiting a decrease in survival of 6 orders of magnitude following a UV-B dose of 250 J m
−2
. Complementation of the
phr
mutations in
P. aeruginosa
PAO1 and
P. syringae
pv. syringae FF5 using the cloned
phr
gene from strain PAO1 resulted in a restoration of survival following UV-B irradiation and recovery under photoreactivating conditions. The UV-B survival of the
phr
mutants could also be complemented by the
P. syringae
mutagenic DNA repair determinant
rulAB
. Assays for increases in the frequency of spontaneous rifampin-resistant mutants in UV-B-irradiated strains containing
rulAB
indicated that significant UV-B mutability (up to a 51-fold increase compared to a nonirradiated control strain) occurred even in the wild-type PAO1 background in which
rulAB
only enhanced the UV-B survival by 2-fold under photoreactivating conditions. The frequency of occurrence of spontaneous nalidixic acid-resistant mutants in the PAO1
uvrA
and
uvrA phr
backgrounds complemented with
rulAB
were 3.8 × 10
−5
and 2.1 × 10
−3
, respectively, following a UV-B dose of 1,550 J m
−2
. The construction and characterization of
phr
mutants in the present study will facilitate the determination of the roles of light and dark repair systems in organisms exposed to solar radiation in their natural habitats.
Proline-Rich Peptide from the Coral Pathogen
Vibrio shiloi
That Inhibits Photosynthesis of Zooxanthellae
Authors: Banin Ehud , Khare Sanjay K. , Naider Fred , Rosenberg Eugene 
Published: July 2002

ABSTRACT

The coral-bleaching bacterium
Vibrio shiloi
biosynthesizes and secretes an extracellular peptide, referred to as toxin P, which inhibits photosynthesis of coral symbiotic algae (zooxanthellae). Toxin P was produced during the stationary phase when the bacterium was grown on peptone or Casamino Acids media at 29°C. Glycerol inhibited the production of toxin P. Toxin P was purified to homogeneity, yielding the following 12-residue peptide: PYPVYAPPPVVP (molecular weight, 1,295.54). The structure of toxin P was confirmed by chemical synthesis. In the presence of 12.5 mM NH
4
Cl, pure natural or synthetic toxin P (10 μM) caused a 64% decrease in the photosynthetic quantum yield of zooxanthellae within 5 min. The inhibition was proportional to the toxin P concentration. Toxin P bound avidly to zooxanthellae, such that subsequent addition of NH
4
Cl resulted in rapid inhibition of photosynthesis. When zooxanthellae were incubated in the presence of NH
4
Cl and toxin P, there was a rapid decrease in the pH (pH 7.8 to 7.2) of the bulk liquid, suggesting that toxin P facilitates transport of NH
3
into the cell. It is known that uptake of NH
3
into cells can destroy the pH gradient and block photosynthesis. This mode of action of toxin P can help explain the mechanism of coral bleaching by
V. shiloi
.
Heterogeneity of Shiga Toxin-Producing
Escherichia coli
Strains Isolated from Hemolytic-Uremic Syndrome Patients, Cattle, and Food Samples in Central France
Authors: Pradel Nathalie , Boukhors Karima , Bertin Yolande , Forestier Christiane , Martin Christine , Livrelli Valérie 
Published: July 2002

ABSTRACT

A detailed analysis of the molecular epidemiology of non-O157:H7 Shiga toxin-producing
Escherichia coli
(STEC) was performed by using isolates from sporadic cases of hemolytic-uremic syndrome (HUS), animal reservoirs, and food products. The isolates belonged to the O91 and OX3 serogroups and were collected in the same geographical area over a short period of time. Five typing methods were used; some of these were used to explore potentially mobile elements like the
stx
genes or the plasmids (
stx
2
-restriction fragment length polymorphism [RFLP],
stx
2
gene variant, and plasmid analyses), and others were used to study the whole genome (ribotyping and pulsed-field gel electrophoresis [PFGE]). The techniques revealed that there was great diversity among the O91 and OX3 STEC strains isolated in central France. A close relationship between strains of the same serotype having the same virulence factor pattern was first suggested by ribotyping. However,
stx
2
-RFLP and
stx
2
variant analyses differentiated all but 5 of 21 isolates, and plasmid analysis revealed further heterogeneity; a unique combination of characteristics was obtained for all strains except two O91:H21 isolates from beef. The latter strains were shown by PFGE to be the most closely related isolates, with >96% homology, and hence may be subtypes of the same strain. Overall, our results indicate that the combination of
stx
2
-RFLP,
stx
2
variant, and plasmid profile analyses is as powerful as PFGE for molecular investigation of STEC diversity. Finally, the non-O157:H7 STEC strains isolated from HUS patients were related to but not identical to those isolated from cattle and food samples in the same geographical area. The possibility that there are distinct lineages of non-O157:H7 STEC, some of which are more virulent for humans, should be investigated further.
Anaerobic Mineralization of Stable-Isotope-Labeled 2-Methylnaphthalene
Authors: Sullivan Elise R. , Zhang Xiaoming , Phelps Craig , Young L. Y. 
Published: July 2002

ABSTRACT

An active sulfate-reducing consortium that degrades 2-methylnaphthalene (2-MNAP) at rates of up to 25 μM day
−1
was established. Degradation was inhibited in the presence of molybdate and ceased in the absence of sulfate. As much as 87% of 2-[
14
C]MNAP was mineralized to
14
CO
2
. 2-Naphthoic acid (2-NA) was detected as a metabolite, and incubation with either deuterated 2-MNAP or [
13
C]bicarbonate indicates that 2-NA is the result of oxidation of the methyl group. Also detected were carboxylated 2-MNAPs, suggesting the presence of an alternative pathway for 2-MNAP degradation.
Method for Spiking Soil Samples with Organic Compounds
Authors: Brinch Ulla C. , Ekelund Flemming , Jacobsen Carsten S. 
Published: July 2002

ABSTRACTWe examined the harmful side effects on indigenous soil microorganisms of two organic solvents, acetone and dichloromethane, that are normally used for spiking of soil with polycyclic aromatic hydrocarbons for experimental purposes. The solvents were applied in two contamination protocols to either the whole soil sample or 25% of the soil volume, which was subsequently mixed with 75% untreated soil. For dichloromethane, we included a third protocol, which involved application to 80% of the soil volume with or without phenanthrene and introduction ofPseudomonas fluorescensVKI171 SJ132 genetically tagged withluxAB::Tn5. For both solvents, application to the whole sample resulted in severe side effects on both indigenous protozoa and bacteria. Application of dichloromethane to the whole soil volume immediately reduced the number of protozoa to below the detection limit. In one of the soils, the protozoan population was able to recover to the initial level within 2 weeks, in terms of numbers of protozoa; protozoan diversity, however, remained low. In soil spiked with dichloromethane with or without phenanthrene, the introducedP. fluorescensVKI171 SJ132 was able to grow to a density 1,000-fold higher than in control soil, probably due mainly to release of predation from indigenous protozoa. In order to minimize solvent effects on indigenous soil microorganisms when spiking native soil samples with compounds having a low water solubility, we propose a common protocol in which the contaminant dissolved in acetone is added to 25% of the soil sample, followed by evaporation of the solvent and mixing with the remaining 75% of the soil sample.
Evaluation of the Abbott AxSYM Cytomegalovirus (CMV) Immunoglobulin M (IgM) Assay in Conjunction with Other CMV IgM Tests and a CMV IgG Avidity Assay
Authors: Lazzarotto T. , Galli C. , Pulvirenti R. , Rescaldani R. , Vezzo R. , La Gioia A. , Martinelli C. , La Rocca S. , Agresti G. , Grillner L. , Nordin M. , van Ranst M. , Combs B. , Maine G. T. , Landini M. P. 
Published: July 2002

ABSTRACT
The measurement of the avidity of cytomegalovirus (CMV) immunoglobulin G (IgG) antibodies has been shown by several investigators to be useful in identifying and excluding primary CMV infections in pregnant women. In this work, we examined the diagnostic utility of reflex testing of CMV IgM-positive specimens from pregnant women by using a CMV IgG avidity assay. The utility of this approach was directly dependent on the sensitivity of the CMV IgM assay employed during the initial screen. The higher initial reactivity rate of the AxSYM CMV IgM assay was necessary in order to detect CMV IgM in specimens containing low-avidity CMV IgG antibodies, indicative of a primary CMV infection, which other CMV IgM assays (Behring, Vidas, Captia, and Eurogenetics) fail to detect in some cases. The use of the AxSYM CMV IgM assay, followed by an avidity test, should result in more accurate diagnosis of CMV infection in pregnant women.
Syphilis Fast Latex Agglutination Test, a Rapid Confirmatory Test
Authors: Fears Martha B. , Pope Victoria 
Published: July 2002

ABSTRACT

Using 255 serum samples with various reactivities, we evaluated the Syphilis Fast latex agglutination test (Syphilis Fast) against the
Treponema pallidum
particle agglutination test (TP-PA) for confirming a diagnosis of syphilis. We found 98.8% agreement between the Syphilis Fast and the TP-PA. The Syphilis Fast, however, had a couple of advantages over the TP-PA: the test takes only 8 min to perform and produces results that are easy to read. It appears to be a good confirmatory test for syphilis, especially for point-of-care clinics such as prenatal or sexually transmitted disease clinics.
Modulation of Release of Proinflammatory Bacterial Compounds by Antibacterials: Potential Impact on Course of Inflammation and Outcome in Sepsis and Meningitis
Authors: Nau Roland , Eiffert Helmut 
Published: July 2002

SUMMARYSeveral bacterial components (endotoxin, teichoic and lipoteichoic acids, peptidoglycan, DNA, and others) can induce or enhance inflammation and may be directly toxic for eukaryotic cells. Bactericidal antibiotics which inhibit bacterial protein synthesis release smaller quantities of proinflammatory/toxic bacterial compounds than Β-lactams and other cell wall-active drugs. Among the Β-lactams, compounds binding to penicillin-binding protein 2 (PBP-2) release smaller amounts of bacterial substances than antibacterials inhibiting PBP-3. Generally, high antibiotic concentrations (more than 10 times the MIC) induce the release of fewer bacterial proinflammatory/toxic compounds than concentrations close to the MIC. In several in vitro and in vivo systems, bacteria treated with protein synthesis inhibitors or Β-lactams inhibiting PBP-2 induce less inflammation than bacteria treated with PBP-3-active Β-lactams. In mouse models of Escherichia coli peritonitis sepsis and of Streptococcus pneumoniae meningitis, lower release of proinflammatory bacterial compounds was associated with reduced mortality. In conclusion, sufficient evidence for the validity of the concept of modulating the release of proinflammatory bacterial compounds by antibacterials has been accumulated in vitro and in animal experiments to justify clinical trials in sepsis and meningitis. A properly conducted study addressing the potential benefit of bacterial protein synthesis inhibitors versus Β-lactam antibiotics will require both strict selection and inclusion of a large number of patients. The benefit of this approach should be greatest in patients with a high bacterial load.
A Postgenomic Approach to Identification ofMycobacterium leprae-Specific Peptides as T-Cell Reagents
Authors: Dockrell Hazel M. , Brahmbhatt Shweta , Robertson Brian D. , Britton Sven , Fruth Uli , Gebre Negussie , Hunegnaw Mesfin , Hussain Rabia , Manandhar Rakesh , Murillo Luis , Pessolani Maria Cristina V. , Roche Paul , Salgado Jorge L. , Sampaio Elizabeth , Shahid Firdaus , Thole Jelle E. R. , Young Douglas B. 
Published: July 2002

ABSTRACTTo identifyMycobacterium leprae-specific human T-cell epitopes, which could be used to distinguish exposure toM. lepraefrom exposure toMycobacterium tuberculosisor to environmental mycobacteria or from immune responses followingMycobacterium bovisBCG vaccination, 15-mer synthetic peptides were synthesized based on data from theM. lepraegenome, each peptide containing three or more predicted HLA-DR binding motifs. Eighty-one peptides from 33 genes were tested for their ability to induce T-cell responses, using peripheral blood mononuclear cells (PBMC) from tuberculoid leprosy patients (n= 59) and healthy leprosy contacts (n= 53) from Brazil, Ethiopia, Nepal, and Pakistan and 20 United Kingdom blood bank donors. Gamma interferon (IFN-γ) secretion proved more sensitive for detection of PBMC responses to peptides than did lymphocyte proliferation. Many of the peptides giving the strongest responses in leprosy donors compared to subjects from the United Kingdom, where leprosy is not endemic, have identical, or almost identical, sequences inM. lepraeandM. tuberculosisand would not be suitable as diagnostic tools. Most of the peptides recognized by United Kingdom donors showed promiscuous recognition by subjects expressing differing HLA-DR types. The majority of the novel T-cell epitopes identified came from proteins not previously recognized as immune targets, many of which are cytosolic enzymes. Fifteen of the tested peptides had ≥5 of 15 amino acid mismatches between the equivalentM. lepraeandM. tuberculosissequences; of these, eight gave specificities of ≥90% (percentage of United Kingdom donors who were nonresponders for IFN-γ secretion), with sensitivities (percentage of responders) ranging from 19 to 47% for tuberculoid leprosy patients and 21 to 64% for healthy leprosy contacts. A pool of such peptides, formulated as a skin test reagent, could be used to monitor exposure to leprosy or as an aid to early diagnosis.
Mutation and Recombination in the Upstream Homology Box-Flanked
ospE
-Related Genes of the Lyme Disease Spirochetes Result in the Development of New Antigenic Variants during Infection
Authors: Sung Shian Ying , McDowell John V. , Carlyon Jason A. , Marconi Richard T. 
Published: July 2002

ABSTRACT

The
ospE
gene family of the Lyme disease spirochetes encodes a polymorphic group of immunogenic lipoproteins. The
ospE
genes are one of several gene families that are flanked by a highly conserved upstream sequence called the upstream homology box, or UHB, element. Earlier analyses in our lab demonstrated that
ospE
-related genes are characterized by defined hypervariable domains (domains 1 and 2) that are predicted to be hydrophilic, surface exposed, and antigenic. The flanking of hypervariable domain 1 by DNA repeats may indicate that recombination contributes to
ospE
diversity and thus ultimately to antigenic variation. Using an isogeneic clone of
Borrelia burgdorferi
B31G (designated B31Gc1), we demonstrate that the
ospE
-related genes undergo mutation and rearrangement during infection in mice. The mutations that develop during infection resulted in the generation of OspE proteins with altered antigenic characteristics. The data support the hypothesized role of OspE-related proteins in immune system evasion.
Induction of Specific Immunoglobulin A and Th2 Immune Responses to P6 Outer Membrane Protein of NontypeableHaemophilus influenzaein Middle Ear Mucosa by Intranasal Immunization
Authors: Kodama Satoru , Suenaga Satoshi , Hirano Takashi , Suzuki Masashi , Mogi Goro 
Published: July 2002

ABSTRACTNontypeableHaemophilus influenzae(NTHI) is a major pathogen of otitis media. One of the outer membrane proteins of NTHI, P6, is an antigen common to all strains and is considered as a candidate for mucosal vaccine. To elucidate the possibility of developing a nasal vaccine against nontypeableHaemophilus influenzae(NTHI) and to investigate mucosal immune responses in the middle ear, mice were immunized intranasally with the P6 outer membrane protein of NTHI, and P6-specific immune responses in the middle ear mucosa were examined. Mice were given with P6 and cholera toxin intranasally as an adjuvant on days 0, 7, and 14 and were killed on day 21. The P6-specific immunoglobulin A (IgA) antibody titer in ear wash was significantly elevated. Mononuclear cells were isolated from middle ear mucosa, and an increase in P6-specific IgA-producing cells was shown with an enzyme-linked immunospot assay. In addition, an increase in memory T cells in middle ear mucosa was detected with flow cytometric analysis after intranasal immunization. Moreover, in vitro stimulation with P6 resulted in proliferation of purified CD4+T cells from immunized mice, and these T cells expressed Th2 cytokine mRNA. These results indicate that P6-specific IgA–B-cell immune responses and selected Th2 cytokine expressing Th cells were induced in middle ear mucosa by intranasal immunization. These findings suggest that a nasal vaccine is useful for preventing otitis media with effusion.
Cryptosporidium parvum
Induces Host Cell Actin Accumulation at the Host-Parasite Interface
Authors: Elliott David A. , Clark Douglas P. 
Published: July 2002

ABSTRACT

Cryptosporidium parvum
is an intracellular protozoan parasite that causes a severe diarrheal illness in humans and animals. Previous ultrastructural studies have shown that
Cryptosporidium
resides in a unique intracellular compartment in the apical region of the host cell. The mechanisms by which
Cryptosporidium
invades host intestinal epithelial cells and establishes this compartment are poorly understood. The parasite is separated from the host cell by a unique electron-dense structure of unknown composition. We have used indirect immunofluorescence microscopy and confocal laser scanning microscopy to characterize this structure. These studies indicate that host filamentous actin is assembled into a plaque-like structure at the host-parasite interface during parasite invasion and persists during parasite development. The actin-binding protein α-actinin is also present in this plaque early in parasite development but is lost as the parasite matures. Other actin-associated proteins, including vinculin, talin, and ezrin, are not present. We have found no evidence of tyrosine phosphorylation within this structure. Molecules known to link actin filaments to membrane were also examined, including α-catenin, β-catenin, plakoglobin, and zyxin, but none was identified at the host-parasite junction. Thus,
Cryptosporidium
induces rearrangement of the host cell cytoskeleton and incorporates host cell actin and α-actinin into a host-parasite junctional complex.
UV-B Irradiation Increases Susceptibility of Mice to Malarial Infection
Authors: Yamamoto Kiichi , Ito Reiko , Koura Minako , Kamiyama Tsuneo 
Published: July 2002

ABSTRACTWe here examined whether exposure of mice to UV-B affected their susceptibility to the murine malaria parasitePlasmodium chabaudi. When BALB/c mice with depilated skin were irradiated with UV-B and subsequently infected with the parasite, 80 to 100% of the UV-B-irradiated mice died within 12 days of infection with a sublethal dose. In addition, UV-B irradiation of C57BL/10 (B-10) mice, which are otherwise naturally resistant to the parasites, rendered them susceptible, and 100% of irradiated B-10 mice died within 11 days postinfection. The level of plasma gamma interferon (IFN-γ) in unirradiated B-10 mice at 5 days after infection increased to 566 pg/ml, whereas the UV-B exposure of mice impaired the production of IFN-γ, which showed a maximum level of 65 pg/ml in response to the parasite infection. The maximum level of plasma interleukin-10 in UV-B-irradiated mice in response to the parasite infection was ∼1,100 pg/ml, which was approximately fourfold higher than the maximum level in unirradiated control mice. When UV-B-irradiated B-10 mice were administered murine recombinant IFN-γ after infection, the mice regained parasite resistance. These results demonstrated that the UV-B exposure of mice enhances the susceptibility to the malaria parasites and suggested that the enhanced susceptibility following UV-B exposure was mediated by impairment of IFN-γ production in response to the parasite infection.
In Vivo-Induced Genes in
Pseudomonas aeruginosa
Authors: Handfield Martin , Lehoux Dario E. , Sanschagrin François , Mahan Michael J. , Woods Donald E. , Levesque Roger C. 
Published: July 2002

ABSTRACT

In vivo expression technology was used for testing
Pseudomonas aeruginosa
in the rat lung model of chronic infection and in a mouse model of systemic infection. Three of the eight
ivi
proteins found showed sequence identity to known virulence factors involved in iron acquisition via an open reading frame (called
pvdI
) implicated in pyoverdine biosynthesis, membrane biogenesis (FtsY), and adhesion (Hag2).
The Response Regulator PhoP Is Important for Survival under Conditions of Macrophage-Induced Stress and Virulence in
Yersinia pestis
Authors: Oyston Petra C. F. , Dorrell Nick , Williams Kerstin , Li Shu-Rui , Green Michael , Titball Richard W. , Wren Brendan W. 
Published: July 2002

ABSTRACT

The two-component regulatory system PhoPQ has been identified in many bacterial species. However, the role of PhoPQ in regulating virulence gene expression in pathogenic bacteria has been characterized only in
Salmonella
species. We have identified, cloned, and sequenced PhoP orthologues from
Yersinia pestis
,
Yersinia pseudotuberculosis
, and
Yersinia enterocolitica
. To investigate the role of PhoP in the pathogenicity of
Y. pestis
, an isogenic
phoP
mutant was constructed by using a reverse-genetics PCR-based strategy. The protein profiles of the wild-type and
phoP
mutant strains, grown at either 28 or 37°C, revealed more than 20 differences, indicating that PhoP has pleiotrophic effects on gene expression in
Y. pestis
. The mutant showed a reduced ability to survive in J774 macrophage cell cultures and under conditions of low pH and oxidative stress in vitro. The mean lethal dose of the
phoP
mutant in mice was increased 75-fold in comparison with that of the wild-type strain, indicating that the PhoPQ system plays a key role in regulating the virulence of
Y. pestis
.