bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a Novel Fimbrial Gene Cluster Related to Long Polar Fimbriae in Locus of Enterocyte Effacement-Negative Strains of Enterohemorrhagic
Escherichia coli
Authors: Doughty Stephen , Sloan Joan , Bennett-Wood Vicki , Robertson Marcus , Robins-Browne Roy M. , Hartland Elizabeth L. 
Published: November 2002

ABSTRACT

Enterohemorrhagic
Escherichia coli
(EHEC) is a food-borne cause of bloody diarrhea and the hemolytic-uremic syndrome (HUS) in humans. Most strains of EHEC belong to a group of bacterial pathogens that cause distinctive lesions on the host intestine termed attaching-and-effacing (A/E) lesions. A/E strains of EHEC, including the predominant serotype, O157:H7, are responsible for the majority of HUS outbreaks worldwide. However, several serotypes of EHEC are not A/E pathogens because they lack the locus of enterocyte effacement (LEE) pathogenicity island. Nevertheless, such strains have been associated with sporadic cases and small outbreaks of hemorrhagic colitis and HUS. Of these LEE-negative organisms, O113:H21 is one of the most commonly isolated EHEC serotypes in many regions. Clinical isolates of LEE-negative EHEC typically express Shiga toxin 2 and carry an ∼90-kb plasmid that encodes EHEC hemolysin, but in the absence of LEE, little is known about the way in which these pathogens colonize the host intestine. In this study we describe the identification of a novel fimbrial gene cluster related to long polar fimbriae in EHEC O113:H21. This chromosomal region comprises four open reading frames,
lpfA
to
lfpD
, and has the same location in the EHEC O113:H21 genome as O island 154 in the prototype EHEC O157:H7 strain, EDL933. In a survey of EHEC of other serotypes, homologues of
lpfA
O113
were found in 26 of 28 LEE-negative and 8 of 11 non-O157:H7 LEE-positive EHEC strains. Deletion of the putative major fimbrial subunit gene,
lpfA
, from EHEC O113:H21 resulted in decreased adherence of this strain to epithelial cells, suggesting that
lpf
O113
may function as an adhesin in LEE-negative isolates of EHEC.
Identification and Genetic Characterization of PmrA-Regulated Genes and Genes Involved in Polymyxin B Resistance in
Salmonella enterica
Serovar Typhimurium
Authors: Tamayo Rita , Ryan Sara S. , McCoy Andrea J. , Gunn John S. 
Published: November 2002

ABSTRACT

Salmonella enterica
serovar Typhimurium encounters antimicrobial peptides (AP) within the phagosomes of professional phagocytes and at intestinal mucosal surfaces.
Salmonella
serovar Typhimurium utilizes the two-component regulatory system PmrA-PmrB, which is activated in response to the environmental conditions encountered in vivo, to regulate resistance to several AP, including polymyxin B (PM). Random Mu
d
J transposon mutagenesis was used to identify PmrA-PmrB-regulated genes, as well as genetic loci necessary for PM resistance. Three different phenotypic classes of genes were identified: those necessary for PM resistance and regulated by PmrA, those necessary for PM resistance and not regulated by PmrA, and PmrA-regulated genes not required for PM resistance. Loci identified as necessary for PM resistance showed between 6- and 192-fold increased sensitivities to PM, and transposon insertion sites include
surA
,
tolB
, and
gnd
. PmrA-regulated loci identified included
dgoA
and
yibD
and demonstrated 500- and 2,500-fold activation by PmrA, respectively. The role of the identified loci in aminoarabinose modification of lipid A was determined by paper chromatography. The
gnd
mutant demonstrated a loss of aminoarabinose from lipid A, which was suggested to be due to a polar effect on the downstream gene
pmrE
. The remaining PM
s
mutants (
surA
and
tolB
), as well as the two PmrA-regulated gene (
yibD
and
dgoA
) mutants, retained aminoarabinose on lipid A.
yibD
,
dgoA
, and
gnd
(likely affecting
pmrE
) played no role in PmrA-regulated resistance to high iron concentrations, while
surA
and
tolB
mutations grew poorly on high iron media. All PM
s
mutants identified in this study demonstrated a defect in virulence compared to wild-type
Salmonella
serovar Typhimurium when administered orally to mice, while the PmrA-regulated gene (
yibD
and
dgoA
) mutants showed normal virulence in mice. These data broaden our understanding of in vivo gene regulation, lipopolysaccharide modification, and mechanisms of resistance to AP in enteric bacteria.
Enhanced Immunogenicity of a Genetic Chimeric Protein Consisting of Two Virulence Antigens ofStreptococcus mutansand Protection against Infection
Authors: Zhang Ping , Jespersgaard Christina , Lamberty-Mallory Leticia , Katz Jannet , Huang Yan , Hajishengallis George , Michalek Suzanne M. 
Published: November 2002

ABSTRACTThe saliva-binding region (SBR) of the cell surface antigen I/II (AgI/II) and the glucan-binding region (GLU) of the glucosyltransferase enzyme ofStreptococcus mutanshave been implicated in the initial adherence ofS. mutansto saliva-coated tooth surfaces and the subsequent sucrose-dependent accumulation ofS. mutans, respectively. Here, we describe the construction and characterization of a genetic chimeric protein consisting of the two virulence determinants SBR and GLU (SBR-GLU). The effectiveness of this construct in inducing mucosal and systemic immune responses to each virulence determinant following intranasal immunization was compared to that of each antigen alone or an equal mixture of SBR and GLU (SBR+GLU) in a mouse model. Furthermore, the ability of antibodies induced to SBR-GLU to protect againstS. mutansinfection was also investigated. Immunization of mice with the chimeric protein SBR-GLU resulted in significantly enhanced (P< 0.001) levels of serum immunoglobulin G (IgG) anti-SBR antibody activity compared to those in the SBR and SBR+GLU groups. The SBR-GLU-immunized mice also demonstrated a significant (P< 0.05) increase in salivary and vaginal IgA antibody responses to SBR and GLU. Analysis of the serum IgG subclass responses to SBR in mice immunized with SBR alone indicated a mixed IgG1 and IgG2a response. A preferential IgG1 response compared to an IgG2a anti-GLU response was induced in mice immunized with GLU alone. Similarly, a preferential IgG1 response was also induced to SBR when GLU was present in either a mixed or conjugated form. Finally, a significant reduction (P< 0.05) inS. mutanscolonization was observed only in mice immunized with the SBR-GLU chimeric protein. Taken together, our results indicate that the chimeric protein SBR-GLU significantly enhanced mucosal immune responses to SBR and GLU and systemic immune responses to SBR. The ability of SBR-GLU to induce responses effective in protection against colonization ofS. mutanssuggests its potential as a vaccine antigen for dental caries.
Analysis ofSalmonella entericaSerotype-Host Specificity in Calves: Avirulence ofS. entericaSerotype Gallinarum Correlates with Bacterial Dissemination from Mesenteric Lymph Nodes and Persistence In Vivo
Authors: Paulin Susan M. , Watson Patricia R. , Benmore Annette R. , Stevens Mark P. , Jones Philip W. , Villarreal-Ramos Bernardo , Wallis Timothy S. 
Published: November 2002

ABSTRACTHost and bacterial factors that determine whetherSalmonellaserotypes remain restricted to the gastrointestinal tract or penetrate beyond the mucosa and cause systemic disease remain largely undefined. Here, factors influencingSalmonellahost specificity in calves were assessed by characterizing the pathogenesis of different serotypes.Salmonella entericaserotype Dublin was highly virulent intravenously, whereasS. entericaserotype Choleraesuis was moderately virulent. Both serotypes were virulent in calves infected orally. In contrast,S. entericaserotypes Gallinarum and Abortusovis were avirulent by either route. Serotypes Dublin, Gallinarum, and Abortusovis colonized the intestinal tract 24 h after oral inoculation, yet only serotype Dublin was consistently recovered from systemic tissues. Serotypes Dublin and Gallinarum invaded bovine intestines in greater numbers and induced greater enteropathogenic responses than serotypes Choleraesuis and Abortusovis. However, only serotype Dublin was able to persist within the intestinal mucosa, and use of a novel cannulation model demonstrated that serotype Dublin was able to pass through the mesenteric lymph nodes in greater numbers than serotype Gallinarum. Together, these results suggest that initial interactions with the intestinal mucosa do not correlate with host specificity, although persistence within tissues and translocation via efferent lymphatics appear to be crucial for the induction of bovine salmonellosis.
The Abundant Polyadenylated Transcript 2 DNA Sequence of the Pathogenic Protozoan Parasite
Entamoeba histolytica
Represents a Nonautonomous Non-Long-Terminal-Repeat Retrotransposon- Like Element Which Is Absent in the Closely Related Nonpathogenic Species
Entamoeba dispar
Authors: Willhoeft Ute , Buß Heidrun , Tannich Egbert 
Published: November 2002

ABSTRACT

While comparing gene expression in the pathogenic organism
Entamoeba histolytica
and the closely related but nonpathogenic species
Entamoeba dispar,
we discovered that the
E. histolytica
abundant polyadenylated transcript 2 (ehapt2) and corresponding genomic copies are absent in
E. dispar
. Although polyadenylated, ehapt2 does not contain any overt open reading frame. Southern blot and sequence analyses revealed that about 500 copies of ehapt2 genomic elements were present in each cell and that the copies were distributed throughout the ameba genome. The various ehapt2 elements are regularly located in the vicinity of protein-encoding genes, downstream of pyrimidine-rich sequence stretches (40 to 125 bp; CT content, 79.2 to 85.5%), and are flanked by duplicated target sites of variable length. Target site duplications were obviously generated during integration of ehapt2 into the
E. histolytica
genome as one copy of the flanking repeat and the complete ehapt2 element are specifically absent in orthologous
E. dispar
genomic sequences. ehapt2 shares 3′ sequences with EhRLE, a recently identified non-long-terminal-repeat (non-LTR) retrotransposon-like element of
E. histolytica
, which contains a conceptual open reading frame for reverse transcriptase. Thus, ehapt2 has all of the properties of nonautonomous non-LTR retrotransposons. A comparison of various
E. histolytica
isolates suggested that transposition of ehapt2 takes place at a very low frequency as the genomic localization of ehapt2 elements was found to be well conserved. A mobile element such as ehapt2 could be a suitable mechanism to explain the infrequent and late transition of
E. histolytica
from a harmless gut commensal to an invasive pathogen.
Identification of a Fibronectin-Binding Protein from
Staphylococcus epidermidis
Authors: Williams Rachel J. , Henderson Brian , Sharp Lindsay J. , Nair Sean P. 
Published: November 2002

ABSTRACT

Staphylococcus epidermidis
has been reported to bind to a number of host cell extracellular matrix proteins, including fibronectin. Here we report the identification of a fibronectin-binding protein from
S. epidermidis
. A phage display library of
S. epidermidis
genomic DNA was constructed and panned against immobilized fibronectin. A number of phagemid clones containing overlapping inserts were identified, and one of these clones, pSE109FN, contained a 1.4-kb insert. Phage pSE109FN was found to bind to fibronectin but not to collagen, fibrinogen, laminin, or vitronectin. However, pSE109FN also bound to heparin, hyaluronate, and plasminogen, although to a lesser extent than it bound to fibronectin. Analysis of The Institute for Genomic Research
S. epidermidis
genome sequence database revealed a 1.85-kb region within a putative 30.5-kb open reading frame, to which the overlapping DNA inserts contained within the fibronectin-binding phagemids mapped. We have designated the gene encoding the fibronectin-binding domain
embp
. A recombinant protein, Embp32, which encompassed the fibronectin-binding domain of Embp, blocked the binding of
S. epidermidis
, but not the binding of
Staphylococcus aureus
, to fibronectin. In contrast, a recombinant protein, FnBPB[D1-D4], spanning the fibronectin-binding domain of the
S. aureus
fibronectin-binding protein FnBPB, blocked binding of
S. aureus
to fibronectin but had a negligible effect on the binding of
S. epidermidis
.
Immunization with the N-Terminal Portion ofTreponema pallidumRepeat Protein K Attenuates Syphilitic Lesion Development in the Rabbit Model
Authors: Morgan Cecilia A. , Lukehart Sheila A. , Van Voorhis Wesley C. 
Published: November 2002

ABSTRACTWhen used as an immunogen,Treponema pallidumrepeat protein K (TprK) has been shown to attenuate syphilitic lesions upon homologous intradermal challenge in the rabbit model. To further explore this protein as a potential vaccine component, we sought to identify the immunogenic regions of TprK. The abilities of three recombinant peptides encompassing TprK to elicit T- and B-cell responses and to protect against challenge were examined. All three fragments elicited proliferative responses from splenocytes taken from infected rabbits. However, enzyme-linked immunosorbent assays indicated that only fragments 1 and 3 were consistently recognized by antisera from infected rabbits. Each fragment was also used to immunize rabbits that were subsequently challenged intradermally with infectiousT. pallidum. All lesions on unimmunized control rabbits ulcerated and contained treponemes, while the lesions on rabbits immunized with fragment 1 were the least likely to have detectable treponemes (25%) and the least likely to ulcerate (37.5%). The lesions on rabbits immunized with fragment 3 showed intermediate results, and rabbits immunized with fragment 2 were the most likely of all those on immunized rabbits to have detectable treponemes (91.7%) and to ulcerate (66.7%). These results demonstrate that epitopes in fragment 1 are recognized by T cells and antibodies during infection and that immunization with this portion of TprK most effectively attenuates syphilitic lesion development.
Search for Potential Vaccine Candidate Open Reading Frames in theBacillus anthracisVirulence Plasmid pXO1: In Silico and In Vitro Screening
Authors: Ariel N. , Zvi A. , Grosfeld H. , Gat O. , Inbar Y. , Velan B. , Cohen S. , Shafferman A. 
Published: November 2002

ABSTRACTA genomic analysis of theBacillus anthracisvirulence plasmid pXO1, aimed at identifying potential vaccine candidates and virulence-related genes, was carried out. The 143 previously defined open reading frames (ORFs) (R. T. Okinaka, K. Cloud, O. Hampton, A. R. Hoffmaster, K. K. Hill, P. Keim, T. M. Koehler, G. Lamke, S. Kumano, J. Mahillon, D. Manter, Y. Martinez, D. Ricke, R. Svensson, and P. J. Jackson, J. Bacteriol.181:6509-6515, 1999) were subjected to extensive sequence similarity searches (with the nonredundant and unfinished microbial genome databases), as well as motif, cellular location, and domain analyses. A comparative genomics analysis was conducted with the related genomes ofBacillus subtilis,Bacillus halodurans, andBacillus cereusand the pBtoxis plasmid ofBacillus thuringiensisvar. israeliensis. As a result, the percentage of ORFs with clues about their functions increased from ∼30% (as previously reported) to more than 60%. The bioinformatics analysis permitted identification of novel genes with putative relevance for pathogenesis and virulence. Based on our analyses, 11 putative proteins were chosen as targets for functional genomics studies. A rapid and efficient functional screening method was developed, in which PCR-amplified full-length linear DNA products of the selected ORFs were transcribed and directly translated in vitro and their immunogenicities were assessed on the basis of their reactivities with hyperimmune anti-B. anthracisantisera. Of the 11 ORFs selected for analysis, 9 were successfully expressed as full-length polypeptides, and 3 of these were found to be antigenic and to have immunogenic potential. The latter ORFs are currently being evaluated to determine their vaccine potential.
Leishmania donovaniSuppresses Activated Protein 1 and NF-κB Activation in Host Macrophages via Ceramide Generation: Involvement of Extracellular Signal-Regulated Kinase
Authors: Ghosh Sanjukta , Bhattacharyya Sandip , Sirkar Madhumita , Sa Gouri Shankar , Das Tanya , Majumdar Debashis , Roy Syamal , Majumdar Subrata 
Published: November 2002

ABSTRACTIn vitro infection of murine peritoneal macrophages with the protozoanLeishmania donovanihas been found to alter the signaling parameters of the host. The present study indicates that the enhancement of intracellular ceramide level in macrophages after infection is a major event relating to macrophage dysfunction. We have previously demonstrated that increased ceramide synthesis in host macrophages was involved in the dephosphorylation of extracellular signal-regulated kinase (ERK). In the present study, we further show that downregulation of ERK by ceramide was found to be associated with the inhibition of activated protein 1 (AP-1) and NF-κB transactivation. Pharmacological inhibition of ceramide synthesis by Fumonisin B1 restored the induction of AP-1 and NF-κB DNA-binding activities in infected BALB/c macrophages. On the contrary, in the case of macrophages from the leishmaniasis-resistant C.D2 mice,L. donovanifailed to induce sustained ceramide synthesis. Enhanced mitogen-activated protein kinase phosphorylation, AP-1 and NF-κB DNA-binding activity, and the generation of nitric oxide (NO) were observed inL. donovani-infected C.D2 macrophages. ERK activation was necessary for the activation of transcription factors AP-1 and NF-κB, NO generation, and restriction of the parasite burden in the resistant murine host macrophages. Hence, the induction of ceramide synthesis in host macrophages appears to be instrumental and one of the turning points leading to silencing of the macrophage antileishmanial responses.
GapA and CrmA Coexpression Is Essential for
Mycoplasma gallisepticum
Cytadherence and Virulence
Authors: Papazisi L. , Frasca S. , Gladd M. , Liao X. , Yogev D. , Geary S. J. 
Published: November 2002

ABSTRACT

It was previously demonstrated that avirulent
Mycoplasma gallisepticum
strain R
high
(passage 164) is lacking three proteins that are expressed in its virulent progenitor, strain R
low
(passage 15). These proteins were identified as the cytadhesin molecule GapA, the putative cytadhesin-related molecule CrmA, and a component of a high-affinity transporter system, HatA. Complementation of R
high
with wild-type
gapA
restored expression in the transformant (GT5) but did not restore the cytadherence phenotype and maintained avirulence in chickens. These results suggested that CrmA might play an essential role in the
M. gallisepticum
cytadherence process. CrmA is encoded by the second gene in the
gapA
operon and shares significant sequence homology to the
ORF6
gene of
Mycoplasma pneumoniae
, which has been shown to play an accessory role in the cytadherence process. Complementation of R
high
with wild-type
crmA
resulted in the transformant (SDCA) that lacked the cytadherence and virulence phenotype comparable to that found in R
high
and GT5. In contrast, complementation of R
high
with the entire wild-type
gapA
operon resulted in the transformant (GCA1) that restored cytadherence to the level found in wild-type R
low
. In vivo pathogenesis trials revealed that GCA1 had regained virulence, causing airsacculitis in chickens. These results demonstrate that both GapA and CrmA are required for
M. gallisepticum
cytadherence and pathogenesis.
Invasion Activity of aMycobacterium tuberculosisPeptide Presented by theEscherichia coliAIDA Autotransporter
Authors: Casali Nicola , Konieczny Marc , Schmidt M. Alexander , Riley Lee W. 
Published: November 2002

ABSTRACTThemce1Agene ofMycobacterium tuberculosiswas initially identified by its ability to promote uptake ofEscherichia coliinto HeLa cells. It was subsequently shown that this activity was confined to a 58-amino-acid region of the protein. A 72-amino-acid fragment (InvX) incorporating this active peptide was expressed inE. colias a fusion to the AIDA (adhesin involved in diffuse adherence) autotransporter translocator, and its stable expression on the surface of the bacterium was demonstrated. RecombinantE. coliexpressing InvX-AIDA showed extensive association with HeLa cells, and InvX was shown to be sufficient for internalization. Uptake was found to be both microtubule and microfilament dependent and required the Rho family of GTPases. Thus, theE. coliAIDA system facilitated both the qualitative and quantitative analysis of the functional domain of a heterologous protein.
Molecular Characterization of a Serotype O121:H19 Clone, a Distinct Shiga Toxin-Producing Clone of PathogenicEscherichia coli
Authors: Tarr Cheryl L. , Large Teresa M. , Moeller Chris L. , Lacher David W. , Tarr Phillip I. , Acheson David W. , Whittam Thomas S. 
Published: November 2002

ABSTRACTMost illnesses caused by Shiga toxin-producingEscherichia coli(STEC) have been attributed toE. coliserotype O157:H7, but non-O157 STEC infections are now increasingly recognized as public health problems worldwide. The O121:H19 serotype is being isolated more frequently from clinical specimens and has been implicated in one waterborne outbreak. We used multilocus virulence gene profiling, a PCR-based assay, to characterize the virulence gene content of 24 isolates of serotype O121:H19 and nonmotile variants. We also performed multilocus enzyme electrophoresis and multilocus sequencing to establish the clonal relatedness of O121 isolates and to elucidate the relationship of O121 to common STEC clones. The 24 isolates were found to represent a single bacterial clone, as there was no allelic variation across 18 enzyme loci among the isolates. The complete nucleotide sequence of the intimin gene differed by four substitutions from that of the epsilon (Int-ε) allele of O103:H2 strain PMK5. The typical O121 virulence gene profile was similar to the profiles of enterohemorrhagicE. coli(EHEC) clones ofE. coli: it included a Shiga toxin 2 gene (stx2), two genes on the EHEC plasmid (toxBandehxA), and the gene encoding intimin (eae). Despite the similarities, putative virulence genes distributed on O islands—large chromosomal DNA segments present in the O157:H7 genome—were useful for discriminating among STEC serotypes and the O121:H19 clone had a composite profile that was distinct from the profiles of the other major EHEC clones of pathogenicE. coli. On the basis of sequencing analysis with 13 housekeeping genes, the O121:H19 clone did not fall into any of the four classical EHEC and enteropathogenicE. coligroups but instead was closely related to twoeae-negative STEC strains.