bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of Tumor Necrosis Factor Alpha and Interleukin 1β in Jejuna of Volunteers after Experimental Challenge with
Cryptosporidium parvum
Correlates with Exposure but Not with Symptoms
Authors: Robinson Prema , Okhuysen Pablo C. , Chappell Cynthia L. , Lewis Dorothy E. , Shahab Imran , Janecki Andrzej , White A. Clinton 
Published: July 2002

ABSTRACT

Jejunal biopsies from volunteers challenged with
Cryptosporidium parvum
were examined for tumor necrosis factor alpha (TNF-α) and interleukin (IL)-1β mRNA. Postchallenge biopsies from 15 of 28 (54%) volunteers expressed TNF-α; 14% expressed IL-1β. Cytokine expression did not correlate with enteric symptoms, suggesting that TNF-α and IL-1β are not key mediators of diarrhea in human cryptosporidiosis.
Cloning of a Beta-Hemolysin Gene ofBrachyspira(Serpulina)hyodysenteriaeand Its Expression inEscherichia coli
Authors: Hsu Tsungda , Hutto David L. , Minion F. Chris , Zuerner Richard L. , Wannemuehler Michael J. 
Published: July 2002

ABSTRACTBrachyspira(Serpulina)hyodysenteriaeinduces a mucohemorrhagic diarrheal disease in pigs. The production of a beta-hemolysin has been considered a major virulence attribute of this organism. Previous reports have failed to correlate a specific cloned gene sequence with a purified beta-hemolytic protein sequence. Thus, questions still remain concerning the structural gene sequence of the hemolysin. To answer this question unequivocally, the beta-hemolytic toxin was purified from extracts of log-phase spirochetes, and the N-terminal amino acid sequence was determined (K-D-V-V-A-N-Q-L-N-I-S-D-K) and compared with the translated sequences of previously cloned genes,tlyAtotlyC. The lack of homology betweentlyAtotlyCtranslated sequences and the purified beta-hemolytic toxin sequence resulted in the study that is reported here. A degenerate probe was designed based on the N-terminal amino acid sequence of the purified beta-hemolysin and used to screen aB. hyodysenteriaegenomic library. Three overlapping clones were identified, and one was sequenced to reveal an open reading frame coding for a putative 8.93-kDa polypeptide containing the N-terminal sequence of the purified beta-hemolysin. To distinguish this gene from thetlyAtotlyCgenes, it has been designatedhlyA. A hemolysis-negativeEscherichia colistrains containinghlyAwas beta-hemolytic on blood agar media. Also, the hemolytic activity of the recombinant protein had identical protease and lipase sensitivities and electrophoretic mobility to those of nativeB. hyodysenteriaebeta-hemolysin. Based on sequence analysis, the translated protein had a pI of 4.3, an α-helical structure, and a phosphopantetheine binding motif. Hybridization analysis of genomic DNA indicated that thehlyAgene was present inB. hyodysenteriaeandB. intermediabut was not detected inB. innocens,B. pilosicoli, orB. murdochiiunder high-stringency conditions. The location ofhlyAon the chromosomal map was distinct from the locations oftlyA,tlyB, andtlyC.
Recombinant PhpA Protein, a Unique Histidine Motif-Containing Protein fromStreptococcus pneumoniae, Protects Mice against Intranasal Pneumococcal Challenge
Authors: Zhang Ying , Masi Amy W. , Barniak Vicki , Mountzouros Ken , Hostetter Margaret K. , Green Bruce A. 
Published: July 2002

ABSTRACTThe multivalent pneumococcal conjugate vaccine is effective against both systemic disease and otitis media caused by serotypes contained in the vaccine. However, serotypes not covered by the current conjugate vaccine may still cause pneumococcal disease. To address these serotypes and the remaining otitis media due toStreptococcus pneumoniae, we have been evaluating antigenically conserved proteins fromS. pneumoniaeas vaccine candidates. A previous report identified a 20-kDa protein with putative human complement C3-proteolytic activity. By utilizing the publicly released pneumococcal genomic sequences, we found the gene encoding the 20-kDa protein to be part of a putative open reading frame of approximately 2,400 bp. We recombinantly expressed a 79-kDa fragment (rPhpA-79) that contains a repeated HxxHxH motif and evaluated it for vaccine potential. The antibodies elicited by the purified rPhpA-79 protein were cross-reactive to proteins from multiple strains ofS. pneumoniaeand were against surface-exposed epitopes. Immunization with rPhpA-79 protein adjuvanted with monophosphoryl lipid A (for subcutaneous immunization) or a mutant cholera toxin, CT-E29H (for intranasal immunization), protected CBA/N mice against death and bacteremia, as well as reduced nasopharyngeal colonization, following intranasal challenge with a heterologous pneumococcal strain. In contrast, immunization with the 20-kDa portion of the PhpA protein did not protect mice. These results suggest that rPhpA-79 is a potential candidate for use as a vaccine against pneumococcal systemic disease and otitis media.
Gene Cluster for Assembly of Pilus Colonization Factor Antigen III of Enterotoxigenic
Escherichia coli
Authors: Taniguchi Tooru , Akeda Yukihiro , Haba Ayako , Yasuda Yoko , Yamamoto Koichiro , Honda Takeshi , Tochikubo Kunio 
Published: July 2002

ABSTRACT

The assembly of pilus colonization factor antigen III (CFA/III) of enterotoxigenic
Escherichia coli
(ETEC) requires the processing of CFA/III major pilin (CofA) by a prepilin peptidase (CofP), similar to other type IV pilus formation systems. CofA is produced initially as a 26.5-kDa preform pilin (prepilin) and then processed to a 20.5-kDa mature pilin by CofP which is predicted to be localized in the inner membrane. In the present experiment, we determined the nucleotide sequence of the whole region for CFA/III formation and identified a cluster of 14 genes, including
cofA
and
cofP
. Several proteins encoded by
cof
genes were similar to previously described proteins, such as the toxin-coregulated pili of
Vibrio cholerae
and the bundle-forming pili of enteropathogenic
E. coli
. The G+C content of the
cof
gene cluster was 37%, which was significantly lower than the average for the
E. coli
genome (50%). The introduction of a recombinant plasmid containing the
cof
gene cluster into the
E. coli
K-12 strain conferred CFA/III biogenesis and the ability of adhesion to the human colon carcinoma cell line Caco-2. This is the first report of a complete nucleotide sequence of the type IV pili found in human ETEC, and our results provide a useful model for studying the molecular mechanism of CFA/III biogenesis and the role of CFA/III in ETEC infection.
Demonstration of Polysaccharide Capsule in
Campylobacter jejuni
Using Electron Microscopy
Authors: Karlyshev Andrey V. , McCrossan Maria V. , Wren Brendan W. 
Published: July 2002

ABSTRACT

Recently, we reported that
Campylobacter jejuni
, an important gastrointestinal pathogen, has the genetic determinants to produce a capsular polysaccharide (Karlyshev et al., Mol. Microbiol. 35:529–541, 2000). Despite these data, the presence of a capsule in these bacteria has remained controversial. In this study we stain
C. jejuni
cells with the cationic dye Alcian blue and demonstrate for the first time by electron microscopy that
C. jejuni
cells produce a polysaccharide capsule that is retained in the coccoid form but is absent in a
kpsM
mutant.
Passive Intranasal Monoclonal Antibody Prophylaxis against MurinePneumocystis cariniiPneumonia
Authors: Gigliotti Francis , Haidaris Constantine G. , Wright Terry W. , Harmsen Allen G. 
Published: July 2002

ABSTRACTPassive antibody immunoprophylaxis is one method used to protect patients against infection if they are unable to mount an adequate active immune response. Topical application of antibody may be effective against infections at mucosal sites. Using a SCID mouse model ofPneumocystis cariniipneumonia, we were able to demonstrate protection against an airborne challenge withP. cariniiby intranasal administration of antibody. Immunoglobulin M (IgM) monoclonal antibodies to an epitope shared by mouse and humanP. cariniiorganisms reduced organism numbers by more than 99% under the conditions described. An IgG1 switch variant of one of the IgM monoclonal antibodies was also protective. These experiments provide a model for exploring the utility of this approach in protecting at-risk patients from infection withP. carinii.
Gender Is a Major Determinant of the Clinical Evolution and Immune Response in Hamsters Infected withLeishmaniaspp
Authors: Travi Bruno L. , Osorio Yaneth , Melby Peter C. , Chandrasekar Bysani , Arteaga Lourdes , Saravia Nancy G. 
Published: July 2002

ABSTRACTIn regions where leishmaniasis is endemic, clinical disease is usually reported more frequently among males than females. This difference could be due to disparate risks of exposure of males and females, but gender-related differences in the host response to infection may also play a role. Experimental studies of the influence of gender onLeishmaniainfection have not included parasites of the subgenusViannia, which is the most common cause of cutaneous leishmaniasis in the Americas. Mice are not readily susceptible to infection byLeishmania(Viannia) spp., but cutaneous infection of hamsters withL.(V.)panamensisorL.(V.)guyanensisresulted in chronic lesions typical of the human disease caused by these parasites. Strikingly, infection of male hamsters resulted in significantly greater lesion size and severity, an increased rate of dissemination to distant cutaneous sites, and a greater parasite burden in the draining lymph node than infection in female animals. Two lines of evidence indicated this gender-related difference in disease evolution was determined at least in part by the sex hormone status of the animal. First, prepubertal male animals had smaller and/or less severe cutaneous lesions than adult male animals. Second, infection of testosterone-treated female animals resulted in significantly larger lesions than in untreated female animals. The increased severity of disease in male compared to female animals was associated with significantly greater intralesional expression of interleukin-4 (IL-4) (P= 0.04), IL-10 (P= 0.04), and transforming growth factor β (TGF-β) (P< 0.001), cytokines known to promote disease in experimental leishmaniasis. There was a direct correlation between the expression of TGF-β mRNA and lesion size (Spearman's correlation coefficient = 0.873;P< 0.001). These findings demonstrate an inherent risk of increased disease severity in male animals, which is associated with a more permissive immune response.
Characterization of a Novel Type IV Pilus Locus Encoded on the Large Plasmid of Locus of Enterocyte Effacement-Negative Shiga-Toxigenic
Escherichia coli
Strains That Are Virulent for Humans
Authors: Srimanote Potjanee , Paton Adrienne W. , Paton James C. 
Published: July 2002

ABSTRACT

The majority of Shiga-toxigenic
Escherichia coli
(STEC) strains isolated from humans with gastrointestinal disease carry large (approximately 90-kb) plasmids. We have been analyzing the megaplasmid (designated pO113) from an O113:H21 STEC strain (98NK2). This strain lacks the locus for enterocyte effacement (LEE) and yet was responsible for an outbreak of hemolytic uremic syndrome. In the present study, we demonstrate that pO113 carries a novel type IV pilus biosynthesis locus (
pil
) related to those of the IncI plasmids R721, R64, and ColIb9. The pO113
pil
locus consists of 11 closely linked genes (
pilL
through
pilV
) with an additional separately transcribed upstream gene (
pilI
). It directs the expression of long thin pili on the 98NK2 surface and the hemagglutination of guinea pig erythrocytes. We also demonstrate that pO113 can be transferred by conjugation. However, the type IV pilus encoded by pO113 does not appear to be involved in the adherence of 98NK2 to HEp-2 or Hct-8 cells in vitro. Homologues of the pO113
pil
locus were present in several other LEE-negative STEC strains but not in LEE-positive STEC strains belonging to serogroup O26, O111, or O157.
Clearance ofHelicobacter pyloriInfection and Resolution of Postimmunization Gastritis in a Kinetic Study of Prophylactically Immunized Mice
Authors: Garhart Christine A. , Redline Raymond W. , Nedrud John G. , Czinn Steven J. 
Published: July 2002

ABSTRACTPatients infected withHelicobacter pylorimount an immune response which fails to clear the infection and may contribute to disease. Mice can be protected by immunization. To further characterize theH. pylori-mouse model, stomachs of unimmunized or intranasally immunized C57BL/6 mice were quantitatively cultured 3 days and 1, 2, 4, 8, 16, 32, and 52 weeks after challenge withH. pylori. At 3 days and 1 week after challenge, colonization was the same in the immunized and unimmunized mice. By 2 weeks after challenge, the immunized mice had a >2-log decrease in bacterial load, and at all later time points, they either were culture negative or had at least a 2-log decrease in bacterial load. Gastritis in the immunized mice peaked at 1 to 2 weeks after challenge and was characterized by a mixed inflammatory infiltrate and epithelial proliferation centered at the transition between corpus and antrum. By 52 weeks postchallenge, the gastric histology in the immunized mice was not different from that in control unchallenged mice. The unimmunized group began to show a reduction in bacterial load as early as 16 weeks after challenge, and by 52 weeks seven of eight unimmunized mice had developed gastritis and reduced bacterial loads. These results indicate that prophylactic immunization does not prevent colonization byH. pyloribut enables mice to clear the infection or significantly reduce the number of colonizing bacteria. The reduction in bacterial load is associated with gastric inflammation that subsides over time.
Characterization of the Distal Tail Fiber Locus and Determination of the Receptor for Phage AR1, Which Specifically Infects
Escherichia coli
O157:H7
Authors: Yu Sung-Liang , Ko Kai-Liang , Chen Chang-Shi , Chang Yu-Chung , Syu Wan-Jr 
Published: July 2002

ABSTRACT

Phage AR1 is similar to phage T4 in several essential genes but differs in host range. AR1 infects various isolates of
Escherichia coli
O157:H7 but does not infect K-12 strains that are commonly infected by T4. We report here the determinants that confer this infection specificity. In T-even phages, gp37 and gp38 are components of the tail fiber that are critical for phage-host interaction. The counterparts in AR1 may be similarly important and, therefore, were characterized. The AR1 gp37 has a sequence that differs totally from those of T2 and T4, except for a short stretch at the N terminus. The gp38 sequence, however, has some conservation between AR1 and T2 but not between AR1 and T4. The sequences that are most closely related to the AR1 gp37 and gp38 are those of phage Ac3 in the T2 family. To identify the AR1-specific receptor,
E. coli
O157:H7 was mutated by Tn
10
insertion and selected for an AR1-resistant phenotype. A mutant so obtained has an insertion occurring at
ompC
that encodes an outer membrane porin. To confirm the role of OmpC in the AR1 infection, homologous replacement was used to create an
ompC
disruption mutant (RM). When RM was complemented with OmpC originated from an O157:H7 strain, but not from K-12, its AR1 susceptibility was fully restored. Our results suggest that the host specificity of AR1 is mediated at least in part through the OmpC molecule.
Insertional Inactivation of Genes Encoding Components of the Sodium-Type Flagellar Motor and Switch of
Vibrio parahaemolyticus
Authors: Boles Blaise R. , McCarter Linda L. 
Published: July 2002

ABSTRACT

Vibrio parahaemolyticus
possesses two types of flagella, polar and lateral, powered by distinct energy sources, which are derived from the sodium and proton motive forces, respectively. Although proton-powered flagella in
Escherichia coli
and
Salmonella enterica
serovar Typhimurium have been extensively studied, the mechanism of torque generation is still not understood. Molecular knowledge of the structure of the sodium-driven motor is only now being developed. In this work, we identify the switch components, FliG, FliM, and FliN, of the sodium-type motor. This brings the total number of genes identified as pertinent to polar motor function to seven. Both FliM and FliN possess charged domains not found in proton-type homologs; however, they can interact with the proton-type motor of
E. coli
to a limited extent. Residues known to be critical for torque generation in the proton-type motor are conserved in the sodium-type motor, suggesting a common mechanism for energy transfer at the rotor-stator interface regardless of the driving force powering rotation. Mutants representing a complete panel of insertionally inactivated switch and motor genes were constructed. All of these mutants were defective in sodium-driven swimming motility. Alkaline phosphatase could be fused to the C termini of MotB and MotY without abolishing motility, whereas deletion of the unusual, highly charged C-terminal domain of FliM disrupted motor function. All of the mutants retained proton-driven, lateral motility over surfaces. Thus, although central chemotaxis genes are shared by the polar and lateral systems, genes encoding the switch components, as well as the motor genes, are distinct for each motility system.
TraG from RP4 and TraG and VirD4 from Ti Plasmids Confer Relaxosome Specificity to the Conjugal Transfer System of pTiC58
Authors: Hamilton Claire M. , Lee Hyewon , Li Pei-Li , Cook David M. , Piper Kevin R. , von Bodman Susanne Beck , Lanka Erich , Ream Walt , Farrand Stephen K. 
Published: July 2002

ABSTRACT

Plasmid conjugation systems are composed of two components, the DNA transfer and replication system, or Dtr, and the mating pair formation system, or Mpf. During conjugal transfer an essential factor, called the coupling protein, is thought to interface the Dtr, in the form of the relaxosome, with the Mpf, in the form of the mating bridge. These proteins, such as TraG from the IncP1 plasmid RP4 (TraG
RP4
) and TraG and VirD4 from the conjugal transfer and T-DNA transfer systems of Ti plasmids, are believed to dictate specificity of the interactions that can occur between different Dtr and Mpf components. The Ti plasmids of
Agrobacterium tumefaciens
do not mobilize vectors containing the
oriT
of RP4, but these IncP1 plasmid derivatives lack the
trans
-acting Dtr functions and TraG
RP4
.
A. tumefaciens
donors transferred a chimeric plasmid that contains the
oriT
and Dtr genes of RP4 and the Mpf genes of pTiC58, indicating that the Ti plasmid mating bridge can interact with the RP4 relaxosome. However, the Ti plasmid did not mobilize transfer from an IncQ relaxosome. The Ti plasmid did mobilize such plasmids if TraG
RP4
was expressed in the donors. Mutations in
traG
RP4
with defined effects on the RP4 transfer system exhibited similar phenotypes for Ti plasmid-mediated mobilization of the IncQ vector. When provided with VirD4, the
tra
system of pTiC58 mobilized plasmids from the IncQ relaxosome. However, neither TraG
RP4
nor VirD4 restored transfer to a
traG
mutant of the Ti plasmid. VirD4 also failed to complement a
traG
RP4
mutant for transfer from the RP4 relaxosome or for RP4-mediated mobilization from the IncQ relaxosome. TraG
RP4
-mediated mobilization of the IncQ plasmid by pTiC58 did not inhibit Ti plasmid transfer, suggesting that the relaxosomes of the two plasmids do not compete for the same mating bridge. We conclude that TraG
RP4
and VirD4 couples the IncQ but not the Ti plasmid relaxosome to the Ti plasmid mating bridge. However, VirD4 cannot couple the IncP1 or the IncQ relaxosome to the RP4 mating bridge. These results support a model in which the coupling proteins specify the interactions between Dtr and Mpf components of mating systems.
The Alkane Hydroxylase Gene of
Burkholderia cepacia
RR10 Is under Catabolite Repression Control
Authors: Marı́n Mercedes M. , Smits Theo H. M. , van Beilen Jan B. , Rojo Fernando 
Published: July 2002

ABSTRACT

In many microorganisms the first step for alkane degradation is the terminal oxidation of the molecule by an alkane hydroxylase. We report the characterization of a gene coding for an alkane hydroxylase in a
Burkholderia cepacia
strain isolated from an oil-contaminated site. The protein encoded showed similarity to other known or predicted bacterial alkane hydroxylases, although it clustered on a separate branch together with the predicted alkane hydroxylase of a
Mycobacterium tuberculosis
strain. Introduction of the cloned
B. cepacia
gene into an alkane hydroxylase knockout mutant of
Pseudomonas fluorescens
CHAO restored its ability to grow on alkanes, which confirms that the gene analyzed encodes a functional alkane hydroxylase. The gene, which was named
alkB
, is not linked to other genes of the alkane oxidation pathway. Its promoter was identified, and its expression was analyzed under different growth conditions. Transcription was induced by alkanes of chain lengths containing 12 to at least 30 carbon atoms as well as by alkanols. Although the gene was efficiently expressed during exponential growth, transcription increased about fivefold when cells approached stationary phase, a characteristic not shared by the few alkane degraders whose regulation has been studied. Expression of the
alkB
gene was under carbon catabolite repression when cells were cultured in the presence of several organic acids and sugars or in a complex (rich) medium. The catabolic repression process showed several characteristics that are clearly different from what has been observed in other alkane degradation pathways.
Two Membrane-Associated NiFeS-Carbon Monoxide Dehydrogenases from the Anaerobic Carbon-Monoxide-Utilizing Eubacterium
Carboxydothermus hydrogenoformans
Authors: Svetlitchnyi Vitali , Peschel Christine , Acker Georg , Meyer Ortwin 
Published: July 2002

ABSTRACT

Two monofunctional NiFeS carbon monoxide (CO) dehydrogenases, designated CODH I and CODH II, were purified to homogeneity from the anaerobic CO-utilizing eubacterium
Carboxydothermus hydrogenoformans
. Both enzymes differ in their subunit molecular masses, N-terminal sequences, peptide maps, and immunological reactivities. Immunogold labeling of ultrathin sections revealed both CODHs in association with the inner aspect of the cytoplasmic membrane. Both enzymes catalyze the reaction CO + H
2
O → CO
2
+ 2 e
−
+ 2 H
+
. Oxidized viologen dyes are effective electron acceptors. The specific enzyme activities were 15,756 (CODH I) and 13,828 (CODH II) μmol of CO oxidized min
−1
mg
−1
of protein (methyl viologen, pH 8.0, 70°C). The two enzymes oxidize CO very efficiently, as indicated by

k
cat
/K
m

values at 70°C of 1.3 · 10
9
M
−1
CO s
−1
(CODH I) and 1.7 · 10
9
M
−1
CO s
−1
(CODH II). The apparent

K
m

values at pH 8.0 and 70°C are 30 and 18 μM CO for CODH I and CODH II, respectively. Acetyl coenzyme A synthase activity is not associated with the enzymes. CODH I (125 kDa, 62.5-kDa subunit) and CODH II (129 kDa, 64.5-kDa subunit) are homodimers containing 1.3 to 1.4 and 1.7 atoms of Ni, 20 to 22 and 20 to 24 atoms of Fe, and 22 and 19 atoms of acid-labile sulfur, respectively. Electron paramagnetic resonance (EPR) spectroscopy revealed signals indicative of [4Fe-4S] clusters. Ni was EPR silent under any conditions tested. It is proposed that CODH I is involved in energy generation and that CODH II serves in anabolic functions.