bluuebunny/crossref_metadata_2025 huggingface dataset:
Microbial Growth on Dichlorobiphenyls Chlorinated on Both Rings as a Sole Carbon and Energy Source
Authors: Kim Sanggoo , Picardal Flynn
Published: July 2002

We have isolated bacterial strains capable of aerobic growth on
ortho
-substituted dichlorobiphenyls as sole carbon and energy sources. During growth on 2,2′-dichlorobiphenyl and 2,4′-dichlorobiphenyl strain SK-4 produced stoichiometric amounts of 2-chlorobenzoate and 4-chlorobenzoate, respectively. Chlorobenzoates were not produced when strain SK-3 was grown on 2,4′-dichlorobiphenyl.
Altered Protein Expression of
Streptococcus oralis
Cultured at Low pH Revealed by Two-Dimensional Gel Electrophoresis
Authors: Wilkins Joanna C. , Homer Karen A. , Beighton David
Published: July 2002

Streptococcus oralis
is the predominant aciduric nonmutans streptococcus isolated from the human dentition, but the role of this organism in the initiation and progression of dental caries has yet to be established. To identify proteins that are differentially expressed by
S. oralis
growing under conditions of low pH, soluble cellular proteins extracted from bacteria grown in batch culture at pH 5.2 or 7.0 were analyzed by two-dimensional (2-D) gel electrophoresis. Thirty-nine proteins had altered expression at low pH; these were excised, digested with trypsin using an in-gel protocol, and further analyzed by peptide mass fingerprinting using matrix-assisted laser desorption ionization mass spectrometry. The resulting fingerprints were compared with the genomic database for
Streptococcus pneumoniae
, an organism that is phylogenetically closely related to
S. oralis
, and putative functions for the majority of these proteins were determined on the basis of functional homology. Twenty-eight proteins were up-regulated following growth at pH 5.2; these included enzymes of the glycolytic pathway (glyceraldehyde-3-phosphate dehydrogenase and lactate dehydrogenase), the polypeptide chains comprising ATP synthase, and proteins that are considered to play a role in the general stress response of bacteria, including the 60-kDa chaperone, Hsp33, and superoxide dismutase, and three distinct ABC transporters. These data identify, for the first time, gene products that may be important in the survival and proliferation of nonmutans aciduric
S. oralis
under conditions of low pH that are likely to be encountered by this organism in vivo.
Seasonal and Spatial Variability in Lake Michigan Sediment Small-Subunit rRNA Concentrations
Authors: MacGregor Barbara J. , Moser Duane P. , Baker Brett J. , Alm Elizabeth W. , Maurer Max , Nealson Kenneth H. , Stahl David A.
Published: July 2002

We have used molecular biological methods to study the distribution of microbial small-subunit rRNAs (SSU rRNAs), in relation to chemical profiles, in offshore Lake Michigan sediments. The sampling site is at a depth of 100 m, with temperatures of 2 to 4°C year-round. RNA extracted from sediment was probed with radiolabeled oligonucleotides targeting bacterial, archaeal, and eukaryotic SSU rRNAs, as well as with a universal probe. The coverage of these probes in relation to the present sequence database is discussed. Because ribosome production is growth rate regulated, rRNA concentrations are an indicator of the microbial populations active in situ. Over a 1-year period, changes in sedimentary SSU rRNA concentrations followed seasonal changes in surface water temperature and SSU rRNA concentration. Sedimentary depth profiles of oxygen, reduced manganese and iron, and sulfate changed relatively little from season to season, but the nitrate concentration was approximately fivefold higher in April and June 1997 than at the other times sampling was done. We propose that sediment microbial SSU rRNA concentrations at our sampling site are influenced by seasonal inputs from the water column, particularly the settling of the spring diatom bloom, and that the timing of this input may be modulated by grazers, such that ammonia becomes available to sediment microbes sooner than fresh organic carbon. Nitrate production from ammonia by autotrophic nitrifying bacteria, combined with low activity of heterotrophic denitrifying bacteria in the absence of readily degradable organic carbon, could account for the cooccurrence of high nitrate and low SSU rRNA concentrations.
Cloning and Expression of an Oligopeptidase, PepO, with Novel Specificity from
Lactobacillus rhamnosus
HN001 (DR20)
Authors: Christensson Camilla , Bratt Henrik , Collins Lesley J. , Coolbear Tim , Holland Ross , Lubbers Mark W. , O’Toole Paul W. , Reid Julian R.
Published: July 2002

Oligopeptidases of starter and nonstarter lactic acid bacteria contribute to the proteolytic events important in maturation and flavor development processes in cheese. This paper describes the molecular cloning, expression, and specificity of the oligopeptidase PepO from the probiotic nonstarter strain
Lactobacillus rhamnosus
HN001 (DR20). The
pepO
gene encodes a protein of 70.9 kDa, whose primary sequence includes the HEXXH motif present in certain classes of metallo-oligopeptidases. The
pepO
gene was cloned in
L. rhamnosus
HN001 and overexpressed in pTRKH2 from its own promoter, which was mapped by primer extension. It was further cloned in both pNZ8020 and pNZ8037 and overexpressed in
Lactococcus lactis
subsp.
cremoris
NZ9000 from the
nisA
promoter. The purified PepO enzyme demonstrated unique cleavage specificity for α
s1
-casein fragment 1–23, hydrolyzing the bonds Pro-5-Ile-6, Lys-7-His-8, His-8-Gln-9, and Gln-9-Gly-10. The impact of this enzyme in cheese can now be assessed.
Semiautomated Quantification of Hepatitis B Virus DNA in a Routine Diagnostic Laboratory
Authors: Kessler Harald H. , Stelzl Evelyn , Daghofer Elisabeth , Santner Brigitte I. , Marth Egon , Lackner Herwig , Stauber Rudolf E.
Published: July 2002

The Cobas Amplicor HBV Monitor test for quantitative determination of hepatitis B virus (HBV) DNA in serum has recently been introduced. To evaluate the performance of this assay in a routine diagnostic laboratory, reproducibility of results was determined with the First European Union Concerted Action HBV Proficiency Panel and the Accurun 325 HBV DNA Positive Control, Series 300. Results for 270 routine serum samples were additionally evaluated. To avoid the retesting of a large number of samples due to titers exceeding the upper limit for the linear range of the assay, sera of patients with chronic hepatitis B (CHB) were diluted prior to the assay to 10
−4
in normal human plasma, which is included in the assay. The mean coefficient of variation was 22.9% for all input HBV DNAs. Of 270 routine serum samples, 182 (150 sera from transplant donors and 32 sera from patients who had recovered from CHB) tested negative. Eighty-six sera were found to be HBV DNA positive; in six sera, HBV DNA levels were found to exceed the upper limit for the linear range of the assay and had to be retested. In the remaining two sera, inhibition occurred. The semiautomated Cobas Amplicor HBV Monitor test showed sufficient reproducibility and helped in avoiding human error. The relatively narrow linear range of detection is a limitation of the new assay.
Dose-Dependent Circulating Immunoglobulin A Antibody-Secreting Cell and Serum Antibody Responses in Swedish Volunteers to an Oral Inactivated Enterotoxigenic
Escherichia coli
Vaccine
Authors: Jertborn Marianne , Åhrén Christina , Svennerholm Ann-Mari
Published: July 2002

The immunogenicity of different preparations of an oral inactivated enterotoxigenic
Escherichia coli
(ETEC) vaccine was evaluated in Swedish volunteers previously unexposed to ETEC infection. The vaccine preparations consisted of recombinant cholera toxin B subunit (CTB) and various amounts of formalin-killed whole bacteria expressing the most prevalent colonization factor antigens (CFAs). Significant immunoglobulin A (IgA) antibody-secreting cell (ASC) responses against CTB and the various CFA components were seen in a majority of volunteers after two doses of ETEC vaccine independent of the vaccine lot given. The IgA ASC responses against CTB were significantly higher after the second than after the first immunization, whereas the CFA-specific IgA ASC responses were almost comparable after the first and second doses of ETEC vaccine. Two immunizations with one-third of a full dose of CFA-ETEC bacteria induced lower frequencies of IgA ASC responses against all the different CFAs than two full vaccine doses, i.e., 63 versus 80% for CFA/I, 56 versus 70% for CS1, 31 versus 65% for CS2, and 56 versus 75% for CS4. The proportion of vaccinees responding with rises in the titer of serum IgA antibody against the various CFA antigens was also lower after immunization with the reduced dose of CFA-ETEC bacteria. These findings suggest that measurements of circulating IgA ASCs can be used not only for qualitative but also for quantitative assessments of the immunogenicity of individual fimbrial antigens in various preparations of ETEC vaccine.
Detection of Serum Thermolabile β-2 Macroglycoprotein (Hakata Antigen) by Enzyme-Linked Immunosorbent Assay Using Polysaccharide Produced by
Aerococcus viridans
Authors: Tsujimura Mitsushi , Ishida Chuzo , Sagara Yasuko , Miyazaki Takashi , Murakami Koichi , Shiraki Hiroshi , Okochi Kazuo , Maeda Yoshiaki
Published: July 2002

Although a serum thermolabile β-2 macroglycoprotein (TMG) may play a role in host defense as a lectin, little is known of its related physiological functions, mainly due to a lack of appropriate methods for tracing the functions of TMG. We identified a polysaccharide from
Aerococcus viridans
, PSA, which reacts with TMG, and based on this finding, we developed an enzyme-linked immunosorbent assay to trace the functions of TMG. Using ethanol precipitation and DEAE-Sepharose and Sephacryl S-400 column chromatographies, we isolated PSA from cultured medium of
A. viridans
, and it exhibited specific binding against TMG in blood samples. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the isolated PSA showed ladder bands that implied the existence of repeating units composed of
d
-glucose,
N
-acetyl-
d
-glucosamine,
d
-mannose, and
d
-xylose, as confirmed by gas chromatography-mass spectrometry. SDS-PAGE and immunochemical analysis, using rabbit anti-TMG antibody, showed that PSA specifically binds solely to intact serum TMG but not to TMG heated at 56°C for 30 min, a condition under which antigenicity is lost. TMG in serum samples bound to PSA in a dose-dependent manner, and this binding was clearly suppressed by addition of PSA. These observations indicate that PSA is a useful adsorbent to TMG and can be used to develop appropriate methods for tracing the functions of TMG.
Roles of Interleukin-12 and Gamma Interferon in MurineChlamydia pneumoniaeInfection
Authors: Geng Yuemei , Berencsi Klara , Gyulai Zsofia , Valyi-Nagy Tibor , Gonczol Eva , Trinchieri Giorgio
Published: July 2002

BALB/c and strain 129 mice infected intranasally withChlamydia pneumoniaedisplayed a moderate-to-severe inflammation in the lungs and produced interleukin-12 (IL-12), gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), and IL-10, with peak levels on days 1 to 3 postinfection (p.i.), returning to basal levels by day 16 p.i. Anti-IL-12 treatment resulted in less-severe pathological changes but higher bacterial titers on days 3 and 7 p.i. By day 16 p.i., the inflammatory responses of control antibody-treated mice subsided. The bacterial titers of both anti-IL-12- and control antibody-treated mice decreased within 3 weeks to marginally detectable levels. Anti-IL-12 treatment significantly reduced lung IFN-γ production and in vitro spleen cell IFN-γ production in response to eitherC. pneumoniaeor concanavalin A. In γ-irradiated infected mice, cytokine production was delayed, and this delay correlated with high bacterial titers in the lungs. FollowingC. pneumoniaeinfection, 129 mice lacking the IFN-γ receptor α chain gene (G129 mice) produced similar IL-12 levels and exhibited similarly severe pathological changes but had higher bacterial titers than 129 mice. However, by day 45 p.i., bacterial titers became undetectable in both wild-type 129 and G129 mice. Thus, duringC. pneumoniaelung infection, IL-12, more than IFN-γ, plays a role in pulmonary-cell infiltration. IFN-γ and IL-12, acting mostly through its induction of IFN-γ and Th1 responses, play an important role in controlling acuteC. pneumoniaeinfection in the lungs, but eventually all mice control the infection to undetectable levels by IL-12- and IFN-γ-independent mechanisms.
Disparate Requirement for T Cells in Resistance to Mucosal and Acute Systemic Candidiasis
Authors: Jones-Carson Jessica , Vazquez-Torres Andres , Warner Thomas , Balish Edward
Published: July 2002

Although highly susceptible to orogastric candidiasis, T-cell receptor δ- and α-chain knockout mice, deficient in γδ and αβ T cells, respectively, were found to be resistant to disseminated candidiasis of endogenous origin and to acute systemic candidiasis (resulting from intravenous injection).
A Subdominant CD8+Cytotoxic T Lymphocyte (CTL) Epitope from thePlasmodium yoeliiCircumsporozoite Protein Induces CTLs That Eliminate Infected Hepatocytes from Culture
Authors: Franke Eileen D. , Sette Alessandro , Sacci John , Southwood Scott , Corradin Giampietro , Hoffman Stephen L.
Published: July 2002

Previous studies indicated that thePlasmodium yoeliicircumsporozoite protein (PyCSP) 57–70 region elicits T cells capable of eliminating infected hepatocytes in vitro. Herein, we report that the PyCSP58–67 sequence contains an H-2dbinding motif, which binds purified Kdmolecules in vitro with low affinity (3,267 nM) and encodes an H-2d-restricted cytotoxic T lymphocyte (CTL) epitope. Immunization of BALB/c mice with three doses of a multiple antigen peptide (MAP) construct containing four branches of amino acids 57 to 70 linked to a lysine-glycine core [MAP4(PyCSP57–70)] and Lipofectin as the adjuvant induced both T-cell proliferation and a peptide-specific CTL response that was PyCSP59–67 specific, H-2drestricted, and CD8+T cell dependent. Immunization with either DNA encoding the PyCSP or irradiated sporozoites demonstrated that this CTL epitope is subdominant since it is not recognized in the context of whole CSP immunization. The biological relevance of this CTL response was underlined by the demonstration that it could mediate genetically restricted, CD8+- and nitric-oxide-dependent elimination of infected hepatocytes in vitro, as well as partial protection of BALB/c mice against sporozoite challenge. These findings indicate that subdominant epitopes with low major histocompatibility complex affinity can be used to engineer epitope-based vaccines and have implications for the selection of epitopes for subunit-based vaccines.
Do Kinetics of the Humoral Response toCryptococcus neoformansProteins during Murine Cryptococcosis Reflect Outcome?
Authors: Neuville Segolene , Lortholary Olivier , Dromer Francoise
Published: July 2002

The kinetics of the humoral response toCryptococcus neoformansproteins were studied in outbred mice infected with isolate NIH52D. Future nonsurvivors had earlier and stronger (i.e., more bands recognized) humoral responses than survivors. In addition, antibodies to a 56- to 60-kDa membrane antigen and to a 39- to 40-kDa cytosolic antigen were detected more frequently in samples from future nonsurvivors and from survivors, respectively (P< 0.05).
Safety and Immunogenicity of Two Different Lots of the Oral, Killed Enterotoxigenic
Escherichia coli
-Cholera Toxin B Subunit Vaccine in Israeli Young Adults
Authors: Cohen Dani , Orr Nadav , Haim Moti , Ashkenazi Shai , Robin Guy , Green Manfred S. , Ephros Moshe , Sela Tamar , Slepon Raphael , Ashkenazi Isaac , Taylor David N. , Svennerholm Ann-Mari , Eldad Arieh , Shemer Joshua
Published: July 2002

Enterotoxigenic
Escherichia coli
(ETEC) is one of the leading causes of diarrhea among Israeli soldiers serving in field units. Two double-blind placebo-controlled, randomized trials were performed among 155 healthy volunteers to evaluate the safety and immunogenicity of different lots of the oral, killed ETEC vaccine consisting of two doses of whole cells plus recombinantly produced cholera toxin B subunit (rCTB). The two doses of vaccine lot E005 and the first dose of vaccine lot E003 were well tolerated by the volunteers. However, 5 (17%) vaccinees reported an episode of vomiting a few hours after the second dose of lot E003; none of the placebo recipients reported similar symptoms. Both lots of vaccine stimulated a rate of significant antibody-secreting cell (ASC) response to CTB and to colonization factor antigen I (CFA/I) after one or two doses, ranging from 85 to 100% and from 81 to 100%, respectively. The rate of ASC response to CS2, CS4, and CS5 was slightly lower than the rate of ASC response induced to CTB, CFA/I, and CS1. The second vaccine dose enhanced the response to CTB but did not increase the frequencies or magnitude of ASC responses to the other antigens. The two lots of the ETEC vaccine induced similar rates of serum antibody responses to CTB and CFA/I which were less frequent than the ASC responses to the same antigens. Based on these safety and immunogenicity data, an efficacy study of the ETEC vaccine is under way in the Israel Defense Force.
Antipeptide Antibody Responses following Intranasal Immunization: Effectiveness of Mucosal Adjuvants
Authors: Olszewska Wieslawa , Partidos Charalambos D. , Steward Michael W.
Published: July 2002

Toxicity is a major factor limiting the development and use of potent adjuvants for human mucosally delivered vaccines. Novel adjuvant formulations have recently become available, and in the present study two have been used for intranasal immunization with a synthetic peptide immunogen (MAP-M2). This peptide represents a multiple antigenic peptide containing multiple copies of a mimotope M2, a peptide mimic of a conformational epitope of the fusion protein of measles virus. MAP-M2 was administered intranasally to experimental animals together with synthetic oligodeoxynucleotides containing unmethylated CpG motifs with or without a mutant of wild-type enterotoxin ofEscherichia coli(LTR72). The combination of the mutant toxin LTR72 and the CpG repeats, codelivered with a peptide immunogen, induced both local and systemic peptide- and pathogen-specific humoral and cellular immune responses comparable to those obtained after intranasal immunization with the wild-type toxin LT. In addition, this combination of adjuvants induced a predominantly immunoglobulin G2a antibody response. If both the LTR72 and CpG adjuvants are shown to be safe for use in humans, this particular combination would appear to have potential as an adjuvant for mucosally delivered vaccines in humans.
Role of
flm
Locus in Mesophilic
Aeromonas
Species Adherence
Authors: Gryllos Ioannis , Shaw Jonathan G. , Gavı́n Rosalina , Merino Susana , Tomás Juan M.
Published: July 2002

The adherence mechanism of
Aeromonas caviae
Sch3N to HEp-2 cells was initially investigated through four mini-Tn
5
mutants that showed a 10-fold decrease in adherence. These mutants lost motility, flagella, and their lipopolysaccharide (LPS) O antigen (O-Ag). Three genes,
flmB-neuA-flmD
, were found to be interrupted by the transposon insertions; additionally, two other genes, one lying upstream (
flmA
) and one downstream (
neuB
), were found to be clustered in the same operon. While the
flmA
and
flmB
genes were present in all mesophilic
Aeromonas
spp. (
A. hydrophila
,
A. caviae
,
A. veronii
bv.
veronii
, and
A. veronii
bv.
sobria
) tested, this was not the case for the
neuA-flmD-neuB
genes. Construction and characterization of
flmB
insertion mutants in five other mesophilic
Aeromonas
strains revealed the loss of motility, flagella, and adherence but did not alter the LPS composition of these strains. Taking the above findings into consideration, we conclude (i) that flagella and possibly the LPS O-Ag are involved in the adherence of the mesophilic
Aeromonas
to human epithelial cells; (ii)
flmA
and
flmB
are genes widely distributed in the mesophilic
Aeromonas
and are involved in flagella assembly, and thus adherence; and (iii) in
A. caviae
Sch3N the
flmA
and
flmB
genes are found in a putative operon together with
neuA
,
flmD
, and
neuB
and are involved in LPS O-Ag biosynthesis and probably have a role in flagellum assembly.
B-Cell Deficiency Suppresses Vaccine-Induced Protection against Murine Filariasis but Does Not Increase the Recovery Rate for Primary Infection
Authors: Martin Coralie , Saeftel Michael , Vuong Phat N. , Babayan Simon , Fischer Kerstin , Bain Odile , Hoerauf Achim
Published: July 2002

To establish the role of B cells and antibodies in destroying filariae, mice lacking mature B cells and therefore unable to produce antibodies were used.Litomosoides sigmodontisoffers a good opportunity for this study because it is the only filarial species that completes its life cycle in mice. Its development was compared in B-cell-deficient mice (BALB/c μMT mice) and wild-type BALB/c mice in two different in vivo situations, vaccination with irradiated larvae and primary infection. In all cases, mice were challenged with subcutaneous inoculation of 40 infective larvae. Vaccine-induced protection was suppressed in B-cell-deficient mice. In these mice, eosinophils infiltrated the subcutaneous tissue normally during immunization; however, their morphological state did not change following challenge inoculation, whereas in wild-type mice the percentage of degranulated eosinophils was markedly increased. From this, it may be deduced that the eosinophil–antibody–B-cell complex is the effector mechanism of protection in vaccinated mice and that its action is fast and takes place in the subcutaneous tissue. In primary infection, the filarial survival and growth was not modified by the absence of B cells. However, no female worm had uterine microfilariae, nor did any mice develop a patent infection. In these mice, concentrations of type 1 (gamma interferon) and type 2 (interleukin-4 [IL-4], IL-5 and IL-10) cytokines in serum were lower and pleural neutrophils were more numerous. The effects of the μMT mutation therefore differ from those in B1-cell-deficient mice described on the same BALB/c background, which reveal a higher filarial recovery rate and microfilaremia. This outlines B2-cell-dependent mechanisms as favorable to the late maturation ofL. sigmodontis.
Reciprocal Immunomodulatory Effects of Gamma Interferon and Interleukin-4 on Filaria-Induced Airway Hyperresponsiveness
Authors: Mehlotra Rajeev K. , Hall Laurie R. , Haxhiu Musa A. , Pearlman Eric
Published: July 2002

Tropical pulmonary eosinophilia (TPE) is a severe asthmatic syndrome of lymphatic filariasis, in which an allergic response is induced to microfilariae (Mf) in the lungs. Previously, in a murine model for TPE, we have demonstrated that recombinant interleukin-12 (IL-12) suppresses pulmonary eosinophilia and airway hyperresponsiveness (AHR) by modulating the T helper (Th) response in the lungs from Th2- to Th1-like, with elevated gamma-interferon (IFN-γ) production and decreased IL-4 and IL-5 production. The present study examined the immunomodulatory roles of IL-4 and IFN-γ in filaria-induced AHR and pulmonary inflammation using mice genetically deficient in these cytokines. C57BL/6, IL-4 gene knockout (IL-4−/−), and IFN-γ−/−mice were first immunized with solubleBrugia malayiantigens and then inoculated intravenously with 200,000 live Mf. Compared with C57BL/6 mice, IL-4−/−mice exhibited significantly reduced AHR, whereas IFN-γ−/−mice had increased AHR. Histopathologically, each mouse strain showed increased cellular infiltration into the lung parenchyma and bronchoalveolar space compared with naı̈ve animals. However, consistent with changes in AHR, IL-4−/−mice had less inflammation than C57BL/6 mice, whereas IFN-γ−/−mice had exacerbated pulmonary inflammation with the loss of pulmonary architecture. Systemically, IL-4−/−mice produced significantly higher IFN-γ levels compared with C57BL/6 mice, whereas IFN-γ−/−mice produced significantly higher IL-4 levels. These data indicate that IL-4 is required for the induction of filaria-induced AHR, whereas IFN-γ suppresses AHR.
DsrA-Deficient Mutant of
Haemophilus ducreyi
Is Impaired in Its Ability To Infect Human Volunteers
Authors: Bong Cliffton T. H. , Throm Robert E. , Fortney Kate R. , Katz Barry P. , Hood Antoinette F. , Elkins Christopher , Spinola Stanley M.
Published: July 2002

Haemophilus ducreyi
produces an outer membrane protein called DsrA, which is required for serum resistance. An isogenic
dsrA
mutant, FX517, was constructed previously in
H. ducreyi
35000. Compared to its parent, FX517 cannot survive in normal human serum. When complemented
in trans
with a plasmid containing
dsrA
, FX517 is converted to a serum-resistant phenotype (C. Elkins, K. J. Morrow, Jr., and B. Olsen, Infect. Immun. 68:1608–1619, 2000). To test whether
dsrA
was transcribed in vivo, we successfully amplified transcripts in five biopsies obtained from four experimentally infected human subjects. To test whether DsrA was required for virulence, six volunteers were experimentally infected with 35000 and FX517 and observed for papule and pustule formation. Each subject was inoculated with two doses (70 to 80 CFU) of live 35000 and 1 dose of heat-killed bacteria on one arm and with three doses (ranging from 35 to 800 CFU) of live FX517 on the other arm. Papules developed at similar rates at sites inoculated with the mutant or parent. However, mutant papule surface areas were significantly smaller than parent papules. The pustule formation rate was 58% (95% confidence interval [CI] of 28 to 85%) at 12 parent sites, and 0% (95% CI of 0 to 15%) at 18 mutant sites (
P
= 0.0004). Although biosafety regulations precluded our testing the complemented mutant in humans, these results suggest that expression of DsrA facilitates the ability of
H. ducreyi
to progress to the pustular stage of disease.
Streptococcus iniaeVirulence Is Associated with a Distinct Genetic Profile
Authors: Fuller Jeffrey D. , Bast Darrin J. , Nizet Victor , Low Donald E. , de Azavedo Joyce C. S.
Published: July 2002

Streptococcus iniaecauses meningoencephalitis and death in commercial fish species and has recently been identified as an emerging human pathogen producing fulminant soft tissue infection. As identified by pulsed-field gel electrophoresis (PFGE), strains causing disease in either fish or humans belong to a single clone, whereas isolates from nondiseased fish are genetically diverse. In this study, we used in vivo and in vitro models to examine the pathogenicity of disease-associated isolates. Strains with the clonal (disease-associated) PFGE profile were found to cause significant weight loss and bacteremia in a mouse model of subcutaneous infection. As little as 102CFU of a disease-associated strain was sufficient to establish bacteremia, with higher inocula (107) resulting in increased mortality. In contrast, non-disease-associated (commensal) strains failed to cause bacteremia and weight loss, even at inocula of 108CFU. In addition, disease-associated strains were more resistant to phagocytic clearance in a human whole blood killing assay compared to commensal strains, which were almost entirely eradicated. Disease-associated strains were also cytotoxic to human endothelial cells as measured by lactate dehydrogenase release from host cells. However, both disease-associated and commensal strains adhered to and invaded cultured human epithelial and endothelial cells equally well. While cellular invasion may still contribute to the pathogenesis of invasiveS. iniaedisease, resistance to phagocytic clearance and direct cytotoxicity appear to be discriminating virulence attributes of the disease-associated clone.
Lack of Expansion of Major Histocompatibility Complex Class Ib-Restricted Effector Cells following Recovery from Secondary Infection with the Intracellular PathogenListeria monocytogenes
Authors: Bouwer H. G. Archie , Barry Ronald A. , Hinrichs David J.
Published: July 2002

Sublethal infection of BALB/c mice with the intracellular bacterial pathogenListeria monocytogenesleads to the development of antilisterial immunity with concurrent stimulation of major histocompatibility complex (MHC) class Ia- and Ib-restricted CD8+effector T cells. SecondaryL. monocytogenesinfection is followed by an accelerated increase in the number ofListeria-specific CD8+cells and rapid clearance of the bacterium from the murine host. Recovery from secondary infection is associated with increased levels of effector cell function, as measured by gamma interferon secretion following coculture of immune cells withL. monocytogenesinfected APCs in vitro, as well as antilisterial cytotoxicity, as measured by effector cell recognition ofL. monocytogenes-infected target cells. We assessed the frequency ofL. monocytogenes-specific MHC class I-restricted cells following secondary infection by ELISPOT assays utilizing coculture of immune cells withL. monocytogenes-infected antigen-presenting cells that express MHC class Ia and/or Ib molecules. We found that the antilisterial Qa-1b(MHC class Ib)-restricted effector subset is not detected as an expanded population following secondary infection compared to the frequency of this effector population as measured following recovery from primary infection. This is in contrast to the frequency of antilisterial H2-Kd(MHC class Ia)-restricted effector cells, which following recovery from secondary infection are detected as an expanded population, and appears to undergo a substantial expansion event 3 to 4 days post-secondary infection. These results are consistent with the conclusion that althoughListeria-specific MHC class Ib-restricted effector cells are present following recovery from secondary infection, this subset does not appear to undergo the expansion phase that is detected for the MHC class Ia-restricted effector cell response.
Antigastric Autoantibodies in Ferrets Naturally Infected with
Helicobacter mustelae
Authors: Cróinı́n Tadhg Ó , Clyne Marguerite , Appelmelk Ben J. , Drumm Brendan
Published: July 2002

Infection with
Helicobacter pylori
has been associated with induction of autoantibodies that cross-react with the gastric mucosa. There have been discordant reports as to whether or not these autoantibodies arise due to molecular mimicry between
H. pylori
and host cell antigens on parietal cells. In this study, we investigated whether molecular mimicry by
H. mustelae
causes autoantibodies in infected ferrets. Serum from
H. mustelae
-infected ferrets reacted with parietal cells in the ferret gastric mucosa but not with duodenal or colonic mucosa. These sera did not react with the blood group A epitope on erythrocytes or
H. mustelae
lipopolysaccharide, and absorption with
H. mustelae
whole cells or red blood cells did not remove autoantibodies. In conclusion, ferrets naturally infected with
H. mustelae
generate antibodies that react with parietal cells, but these autoantibodies are not due to molecular mimicry.
Induction of Systemic Antifimbria and Antitoxin Antibody Responses in Egyptian Children and Adults by an Oral, Killed Enterotoxigenic
Escherichia coli
plus Cholera Toxin B Subunit Vaccine
Authors: Hall Eric R. , Wierzba Thomas F. , Åhrén Christina , Rao Malla R. , Bassily Samir , Francis Wagdy , Girgis Fouad Y. , Safwat Mohamed , Lee Young J. , Svennerholm Ann-Mari , Clemens John D. , Savarino Stephen J.
Published: July 2002

We assessed serologic responses to an oral, killed whole-cell enterotoxigenic
Escherichia coli
plus cholera toxin B-subunit (ETEC-rCTB) vaccine in 73 Egyptian adults, 105 schoolchildren, and 93 preschool children. Each subject received two doses of vaccine or placebo 2 weeks apart, giving blood before immunization and 7 days after each dose. Plasma antibodies to rCTB and four vaccine-shared colonization factors (CFs) were measured by enzyme-linked immunosorbent assay. Immunoglobulin A (IgA) antibodies to rCTB and CFA/I were measured in all subjects, and those against CS1, CS2, and CS4 were measured in all children plus a subset of 33 adults. IgG antibodies to these five antigens were measured in a subset of 30 to 33 subjects in each cohort. Seroconversion was defined as a >2-fold increase in titer after vaccination. IgA and IgG seroconversion to rCTB was observed in 94 to 95% of adult vaccinees, with titer increases as robust as those previously reported for these two pediatric cohorts. The proportion showing IgA seroconversion to each CF antigen among vaccinated children (range, 70 to 96%) and adults (31 to 69%), as well as IgG seroconversion in children (44 to 75%) and adults (25 to 81%), was significantly higher than the corresponding proportion in placebo recipients, except for IgA responses to CS2 in adults. IgA anti-CF titers peaked after one dose in children, whereas in all age groups IgG antibodies rose incrementally after each dose. Independently, both preimmunization IgA titer and age were inversely related to the magnitude of IgA responses. In conclusion, serologic responses to the ETEC-rCTB vaccine may serve as practical immune outcome measures in future pediatric trials in areas where ETEC is endemic.
Characterization of Proteins in the Outer Membrane Preparation of a Murine Pathogen,
Helicobacter bilis
Authors: Ge Zhongming , Doig Peter , Fox James G.
Published: July 2002

Helicobacter bilis
is a bacterial pathogen associated with multifocal hepatitis and inflammatory bowel disease in certain strains of mice. This bacterium colonizes the liver, bile, and lower intestine in mice and has also been isolated from a wide spectrum of laboratory animals. In this study, proteins present in the outer membrane preparation (OMP) of four
H. bilis
strains isolated from a mouse, a dog, a rat, and a gerbil were characterized and compared with that of
Helicobacter pylori
, a human gastric pathogen. All four
H. bilis
strains had similar OMP protein profiles that were distinct from those of
H. pylori
. Immunoblotting demonstrated that OMP proteins from
H. bilis
and
H. pylori
have little cross-reactivity, except for their flagellins. Nine major immunogenic polypeptides were present in the
H. bilis
OMPs. By using two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, five heat-modifiable proteins with molecular masses of 82, 66, 52, 47 and 37 kDa were identified. The N-terminal sequences of the 46- and 47-kDa OMP proteins had no homology with protein sequences available in public databases. These results indicate that
H. bilis
has a conserved, unique OMP protein profile that is distinct from those of
H. pylori
.
Identification of a Novel T-Cell Epitope in Soluble Egg Antigen ofSchistosoma japonicum
Authors: Liu Jie , Tasaka Kachio , Yang Junqi , Itoh Takehito , Yamada Muneo , Yoshikawa Hideshi , Nakajima Yasuo
Published: July 2002

Identification of T-cell epitopes harbored in soluble egg antigen (SEA) ofSchistosoma japonicumand study of the immunological properties are essential for understanding the immunopathology and the control of schistosomiasis. As a follow-up to our previous work, the 66- to 80-kDa fragment from SEA was partially digested with protease, fractionated by reverse-phase high-pressure liquid chromatography, and found to be carrying a peptide which stimulated proliferation and gamma interferon (IFN-γ) production of Th1 clones specific to SEA. Sequence analysis showed that the peptide was composed of 12 amino acids lined up as DLAVELAYLGNL. A synthetic homologue induced proliferation and IFN-γ and interleukin-2 (IL-2) production, but not IL-4 or IL-6 production, by the Th1 clones as well as by the spleen cells from SEA-immunized mice, thus indicating that the peptide carries a Th1 epitope of SEA.
S-Fimbria-Encoding Determinant
sfa
I
Is Located on Pathogenicity Island III
536
of Uropathogenic
Escherichia coli
Strain 536
Authors: Dobrindt Ulrich , Blum-Oehler Gabriele , Hartsch Thomas , Gottschalk Gerhard , Ron Eliora Z. , Fünfstück Reinhard , Hacker Jörg
Published: July 2002

The
sfa
I
determinant encoding the S-fimbrial adhesin of uropathogenic
Escherichia coli
strains was found to be located on a pathogenicity island of uropathogenic
E. coli
strain 536. This pathogenicity island, designated PAI III
536
, is located at 5.6 min of the
E. coli
chromosome and covers a region of at least 37 kb between the tRNA locus
thrW
and
yagU
. As far as it has been determined, PAI III
536
also contains genes which code for components of a putative enterochelin siderophore system of
E. coli
and
Salmonella
spp. as well as for colicin V immunity. Several intact or nonfunctional mobility genes of bacteriophages and insertion sequence elements such as transposases and integrases are present on PAI III
536
. The presence of known PAI III
536
sequences has been investigated in several wild-type
E. coli
isolates. The results demonstrate that the determinants of the members of the S-family of fimbrial adhesins may be located on a common pathogenicity island which, in
E. coli
strain 536, replaces a 40-kb DNA region which represents an
E. coli
K-12-specific genomic island.
Towards Development of an Edible Vaccine against Bovine Pneumonic Pasteurellosis Using Transgenic White Clover Expressing aMannheimia haemolyticaA1 Leukotoxin 50 Fusion Protein
Authors: Lee Raymond W. H. , Strommer Judith , Hodgins Doug , Shewen Patricia E. , Niu Yongqing , Lo Reggie Y. C.
Published: July 2002

Development of vaccines against bovine pneumonia pasteurellosis, or shipping fever, has focused mainly onMannheimia haemolyticaA1 leukotoxin (Lkt). In this study, the feasibility of expressing Lkt in a forage plant for use as an edible vaccine was investigated. Derivatives of theM. haemolyticaLkt in which the hydrophobic transmembrane domains were removed were made. Lkt66 retained its immunogenicity and was capable of eliciting an antibody response in rabbits that recognized and neutralized authentic Lkt. Genes encoding a shorter Lkt derivative, Lkt50, fused to a modified green fluorescent protein (mGFP5), were constructed for plant transformation. Constructs were screened by Western immunoblot analysis for their ability to express the fusion protein after agroinfiltration in tobacco. The fusion construct pBlkt50-mgfp5, which employs the cauliflower mosaic virus 35S promoter for transcription, was selected and introduced into white clover byAgrobacterium tumefaciens-mediated transformation. Transgenic lines of white clover were recovered, and expression of Lkt50-GFP was monitored and confirmed by laser confocal microscopy and Western immunoblot analysis. Lkt50-GFP was found to be stable in clover tissue after drying of the plant material at room temperature for 4 days. An extract containing Lkt50-GFP from white clover was able to induce an immune response in rabbits (via injection), and rabbit antisera recognized and neutralized authentic Lkt. This is the first demonstration of the expression of anM. haemolyticaantigen in plants and paves the way for the development of transgenic plants expressingM. haemolyticaantigens as an edible vaccine against bovine pneumonic pasteurellosis.
Bacterial Lipoprotein-Based Vaccines Induce Tumor Necrosis Factor-Dependent Type 1 Protective Immunity againstLeishmania major
Authors: Cote-Sierra Javier , Bredan Amin , Toldos Carmen M. , Stijlemans Benoit , Brys Lea , Cornelis Pierre , Segovia Manuel , de Baetselier Patrick , Revets Hilde
Published: July 2002

Immunity againstLeishmania majorrequires rapid induction of a type 1 immune response in which tumor necrosis factor alpha (TNF-α) plays an essential role. Hence, vaccination strategies that simulate the protective immune response found in hosts that have recovered from natural infection provide a rational approach to combat leishmaniasis. One method for optimizing the qualitative and quantitative immune responses after vaccination is to use an adjuvant. In this study we demonstrate that the OprI lipoprotein (L-OprI) fromPseudomonas aeruginosainduces a long-term cellular (gamma interferon [IFN-γ]) and humoral (immunoglobulin G2a) type 1 immune response against a truncated 32-kDa version (COOHgp63) of the 63-kDa major cell surface glycoprotein gp63. By contrast, immunization with COOHgp63 either fused to OprI nonlipoprotein or with no adjuvant did not result in the induction of type 1 immune responses. The adjuvanticity of L-OprI is strongly dependent on its capacity to induce TNF-α, since generation of type 1 immune responses is clearly delayed and impaired in TNF-α−/−mice. Vaccination with L-OprICOOHgp63 fusion protein protected BALB/c mice againstL. majorinfection for at least 19 weeks. Vaccinated mice were largely free of lesions or clearly controlled lesion size on termination of the experiment. The control of disease progression in mice vaccinated with L-OprICOOHgp63 was associated with enhancement of antigen-specific IFN-γ production. These data indicate that bacterial lipoproteins constitute appropriate adjuvants to include in vaccines against diseases in which type 1 immune responses are important for protection.
Molecular Analysis of Group B Protective Surface Protein, a New Cell Surface Protective Antigen of Group B Streptococci
Authors: Erdogan Sezgin , Fagan Peter K. , Talay Susanne R. , Rohde Manfred , Ferrieri Patricia , Flores Aurea E. , Guzmán Carlos A. , Walker Mark J. , Chhatwal Gursharan S.
Published: July 2002

Group B streptococci (GBS) express various surface antigens designated c, R, and X antigens. A new R-like surface protein from
Streptococcus agalactiae
strain Compton R has been identified by using a polyclonal antiserum raised against the R protein fraction of this strain to screen a lambda Zap library. DNA sequence analysis of positive clones allowed the prediction of the primary structure of a 105-kDa protein designated BPS protein (group B protective surface protein) that exhibited typical features of streptococcal surface proteins such as a signal sequence and a membrane anchor region but did not show significant similarity with other known sequences. Immunogold electron microscopy using a BPS-specific antiserum confirmed the surface location of BPS protein on
S. agalactiae
strain Compton R. Anti-BPS antibodies did not cross-react with R1 and R4 proteins expressed by two variant type III GBS strains but reacted with the parental streptococcal strain in Western blot and immunoprecipitation analyses. Separate R3 and BPS immunoprecipitation bands were observed when a cell extract of strain Compton R was tested with an antiserum against Compton R previously cross-absorbed to remove R4 antibodies. Immunization of mice with recombinant BPS protein by the subcutaneous route produced an efficient antigen-specific response, and immunized animals survived challenge with a lethal dose of a virulent strain. Therefore, BPS protein represents a new R-like protective antigen of GBS.
Cross-Protective Immunity of Mice Induced by Oral Immunization with Pneumococcal Surface Adhesin A Encapsulated in Microspheres
Authors: Seo Jun-Young , Seong Seung Yong , Ahn Byung-Yoon , Kwon Ick Chan , Chung Hesson , Jeong Seo Young
Published: July 2002

The global use of a capsular polysaccharide-based pneumococcal vaccine has been limited because of serotype-specific protection and poor effectiveness in individuals with low immunocompetency. The mucosal immune system develops earlier in infants and lasts longer in the elderly than does the systemic immune system. Furthermore, mucosal immunization is beneficial for AIDS patients, because human immunodeficiency virus-infected subjects can develop normal mucosal antibody responses even in late clinical phases. For these reasons, we evaluated recombinant pneumococcal surface adhesin A (rPsaA) ofStreptococcus pneumoniaein terms of cross-protective immune responses after oral delivery. Encapsulated rPsaA provided higher immunogenicity than naked rPsaA. Coencapsulation or codelivery of the cholera toxin (CT) B subunit (CTB) and CT also increased the immunogenicity of rPsaA. Cross-protective immunities against five strains ofS. pneumoniae(types 4, 6B, 14, 19F, and 23F) were induced after oral immunization with microencapsulated rPsaA. Lung colonization and septicemia caused by the five serotypes were significantly inhibited by oral immunization with microencapsulated rPsaA. These results suggest that rPsaA coencapsulated with CTB can be used as an oral vaccine to induce cross-protective immunity for the prevention of pneumococcal infection.
Increased Interleukin 4 (IL-4) Receptor Expression and IL-4-Induced Decrease in IL-12 Production by Langerhans Cells Infected withLeishmania major
Authors: Moll Heidrun , Scharner Anabel , Kämpgen Eckhart
Published: July 2002

Langerhans cells (LC) take upLeishmania majorand are critical for the induction of the parasite-specific T-cell response. Their functional activities are regulated by cytokines. We analyzed whether infection of LC withL. majormodulates the expression of their cytokine receptors. The expression of the interleukin 4 (IL-4) receptor was increased on infected LC from susceptible mice but not on those from resistant mice. Moreover, IL-4 treatment strongly decreased the lipopolysaccharide-induced IL-12 response of infected LC from susceptible mice. This modulation of IL-4 receptor expression and IL-12 production by infection of LC withLeishmaniamay contribute to the development of Th2 cells and to susceptibility to infection.
The Major Structural Subunits of Dr and F1845 Fimbriae Are Adhesins
Authors: Van Loy Cristina P. , Sokurenko Evgeni V. , Moseley Steve L.
Published: July 2002

Fimbrial adhesins mediate the attachment of pathogenic
Escherichia coli
to various host tissues leading to the development of disease. The Dr hemagglutinin and F1845 fimbriae belong to the Dr family of adhesins, which is associated with urinary tract infections and diarrheal disease. These adhesins bind to the Dr
a
blood-group antigen present on decay-accelerating factor (DAF). The Dr hemagglutinin is unique in this family since it also binds to type IV collagen and its binding is inhibited by the presence of chloramphenicol. We have purified the major structural subunits of Dr and F1845 fimbriae, DraE and DaaE, as fusions to maltose-binding protein and to oligohistidine tags and examined their binding to erythrocytes, Chinese hamster ovary cell transfectants expressing DAF, and a DAF fusion protein. The DraE and DaaE fusion proteins bind to the DAF receptor in a specific manner resembling the distinct phenotypes of the corresponding Dr and F1845 fimbriae. In contrast to binding studies with the DAF receptor, the DraE fusion proteins did not bind to type IV collagen.