bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel α-Glucosidase from
Aspergillus nidulans
with Strong Transglycosylation Activity
Authors: Kato Naoki , Suyama Sachie , Shirokane Masao , Kato Masashi , Kobayashi Tetsuo , Tsukagoshi Norihiro 
Published: July 2002

ABSTRACT

Aspergillus nidulans
possessed an α-glucosidase with strong transglycosylation activity. The enzyme, designated α-glucosidase B (AgdB), was purified and characterized. AgdB was a heterodimeric protein comprising 74- and 55-kDa subunits and catalyzed hydrolysis of maltose along with formation of isomaltose and panose. Approximately 50% of maltose was converted to isomaltose, panose, and other minor transglycosylation products by AgdB, even at low maltose concentrations. The
agdB
gene was cloned and sequenced. The gene comprised 3,055 bp, interrupted by three short introns, and encoded a polypeptide of 955 amino acids. The deduced amino acid sequence contained the chemically determined N-terminal and internal amino acid sequences of the 74- and 55-kDa subunits. This implies that AgdB is synthesized as a single polypeptide precursor. AgdB showed low but overall sequence homology to α-glucosidases of glycosyl hydrolase family 31. However, AgdB was phylogenetically distinct from any other α-glucosidases. We propose here that AgdB is a novel α-glucosidase with unusually strong transglycosylation activity.
Identification of Quorum-Quenching
N
-Acyl Homoserine Lactonases from
Bacillus
Species
Authors: Dong Yi-Hu , Gusti Andi R. , Zhang Qiong , Xu Jin-Ling , Zhang Lian-Hui 
Published: July 2002

ABSTRACT

A range of gram-negative bacterial species use
N
-acyl homoserine lactone (AHL) molecules as quorum-sensing signals to regulate different biological functions, including production of virulence factors. AHL is also known as an autoinducer. An autoinducer inactivation gene,
aiiA
, coding for an AHL lactonase, was cloned from a bacterial isolate,
Bacillus
sp. strain 240B1. Here we report identification of more than 20 bacterial isolates capable of enzymatic inactivation of AHLs from different sources. Eight isolates showing strong AHL-inactivating enzyme activity were selected for a preliminary taxonomic analysis. Morphological phenotypes and 16S ribosomal DNA sequence analysis indicated that these isolates probably belong to the species
Bacillus thuringiensis.
Enzymatic analysis with known
Bacillus
strains confirmed that all of the strains of
B. thuringiensis
and the closely related species
B. cereus
and
B. mycoides
tested produced AHL-inactivating enzymes but
B. fusiformis
and
B. sphaericus
strains did not. Nine genes coding for AHL inactivation were cloned either by functional cloning or by a PCR procedure from selected bacterial isolates and strains. Sequence comparison of the gene products and motif analysis showed that the gene products belong to the same family of AHL lactonases.
Analysis of
Streptococcus mutans
Proteins Modulated by Culture under Acidic Conditions
Authors: Wilkins Joanna C. , Homer Karen A. , Beighton David 
Published: July 2002

ABSTRACT

Streptococcus mutans
, a major etiological agent of dental caries, causes demineralization of the tooth tissue due to the formation of acids from dietary carbohydrates. Dominant among the virulence determinants of this organism are aciduricity and acidogenicity, the abilities to grow at low pH and to produce acid, respectively. The mechanisms underlying the ability of
S. mutans
to survive and proliferate at low pH are currently under investigation. In this study we cultured
S. mutans
at pH 5.2 or 7.0 and extracted soluble cellular proteins. These were analyzed using high-resolution two-dimensional gel electrophoresis, and replicate maps of proteins expressed under each of the two conditions were generated. Proteins with modulated expression at low pH, as judged by a change in the relative integrated optical density, were excised and digested with trypsin by using an in-gel protocol. Tryptic digests were analyzed using matrix-assisted laser desorption ionization mass spectrometry to generate peptide mass fingerprints, and these were used to assign putative functions according to their homology with the translated sequences in the
S. mutans
genomic database. Thirty individual proteins exhibited altered expression as a result of culture of
S. mutans
at low pH. Up-regulated proteins (
n
= 18) included neutral endopeptidase, phosphoglucomutase, 60-kDa chaperonin, cell division proteins, enolase, lactate dehydrogenase, fructose bisphosphate aldolase, acetoin reductase, superoxide dismutase, and lactoylglutathione lyase. Proteins down-regulated at pH 5.2 (
n
= 12) included protein translation elongation factors G, Tu, and Ts, DnaK, small-subunit ribosomal protein S1P, large-subunit ribosomal protein L12P, and components of both phosphoenolpyruvate:protein phosphotransferase and multiple sugar binding transport systems. The identification of proteins differentially expressed following growth at low pH provides new information regarding the mechanisms of survival and has identified new target genes for mutagenesis studies to further assess their physiological significance.
Comparative Phenotypic, Molecular, and Virulence Characterization of
Vibrio parahaemolyticus
O3:K6 Isolates
Authors: Yeung P. S. Marie , Hayes Micaela C. , DePaola Angelo , Kaysner Charles A. , Kornstein Laura , Boor Kathryn J. 
Published: July 2002

ABSTRACT

Historically,
Vibrio parahaemolyticus
infections have been characterized by sporadic cases caused by multiple, diverse serotypes. However, since 1996,
V. parahaemolyticus
serotype O3:K6 strains have been associated with several large-scale outbreaks of illness, suggesting the emergence of a “new” group of organisms with enhanced virulence. We have applied three different molecular subtyping techniques to identify an appropriate method for differentiating O3:K6 isolates from other serotypes. Pulsed-field gel electrophoresis (PFGE) following
Not
I digestion differentiated seven closely related subtypes among O3:K6 and related strains, which were distinct from PFGE patterns for non-O3:K6 isolates. Ribotyping and
tdh
sequencing were less discriminatory than PFGE, but further confirmed close genetic relationships among recent O3:K6 isolates. In vitro adherence and cytotoxicity studies with human epithelial cells showed that O3:K6 isolates exhibited statistically higher levels of adherence and cytotoxicity to host cells than non-O3:K6 isolates. Epithelial cell cytotoxicity patterns were determined with a lactate dehydrogenase release assay. At 3 h postinfection, high relative cytotoxicities (>50% maximum lactate dehydrogenase activity) were found among a greater proportion of recently isolated O3:K6 and closely related strains (75%) than among the non-O3:K6 isolates (23%). A statistically significant relationship between adherence and cytotoxicity suggests that the pathogenic potential of some isolates may be associated with increased adherence to epithelial cells. Our findings suggest that enhanced adherence and cytotoxicity may contribute to the apparent unique pathogenic potential of
V. parahaemolyticus
O3:K6 strains.
Casein Utilization by
Streptococcus thermophilus
Results in a Diauxic Growth in Milk
Authors: Letort Catherine , Nardi Michèle , Garault Peggy , Monnet Véronique , Juillard Vincent 
Published: July 2002

ABSTRACT

In milk,
Streptococcus thermophilus
displays two distinct exponential growth phases, separated by a nonexponential one, during which proteinase synthesis was initiated. During the second exponential phase, utilization of caseins as the source of amino acids resulted in a decrease in growth rate, presumably caused by a limiting peptide transport activity.
Lipid Removal from Human Serum Samples
Authors: Castro Arnold R. , Morrill William E. , Pope Victoria 
Published: July 2002

ABSTRACTThe efficacy of lipid removal from human serum samples obtained by using Cleanascite HC, a commercially available product, was compared to that obtained by the standard chloroform method. Separate samples of 21 frozen, banked human serum samples used in the preparation of samples for proficiency testing were treated with either Cleanascite HC or chloroform. The lipid content was measured before and after treatment. The total percentages of lipid removed ranged from 61 to 70% with Cleanascite HC and from 60 to 62% with chloroform. The advantage of Cleanascite HC over chloroform is based on the simplicity of the procedure with Cleanascite HC without the environmental concerns inherent in the use of chloroform. In 15 serum samples known to contain antibodies to treponemal and nontreponemal syphilis antigens, Cleanascite HC bound some immunoglobulin, but with only minimal loss of reactivity in the serologic tests for syphilis. Cleanascite HC is therefore an acceptable alternative to chloroform for lipid reduction in human serum samples.
Molecular Markers in Acute and Chronic Phases of Human Toxoplasmosis: Determination of Immunoglobulin G Avidity by Western Blotting
Authors: Marcolino P. T. , Silva D. A. O. , Leser P. G. , Camargo M. E. , Mineo J. R. 
Published: July 2002

ABSTRACT

We characterized antigenic markers recognized by human serum samples from patients presenting with acute and chronic toxoplasmosis by the determination of immunoglobulin G (IgG) antibody avidity by a Western blot modified technique (avidity immunoblotting) that includes the dissociation of the antigen-antibody interaction with 6 or 8 M urea solutions. Human serum samples from 20 patients presenting with recent infection and from 20 patients with chronic infection were analyzed. It was observed that bands p16, p32, p38, p40, p43, p54, p60, p66, and p97 were more frequently recognized by low-avidity IgG in recent infection and by high-avidity IgG in chronic toxoplasmosis. From these antigenic bands, p38 can be characterized as an optimal antigenic marker of low avidity for recent forms of toxoplasmosis due to a significant decrease of their frequencies (from 80 to 0%) after treatment with 6 M urea solutions. The p30 antigen was not considered a good marker to distinguish acute from chronic infection since corresponding IgG antibodies were determined to have high avidity in both phases of the infection. Thus, the avidity immunoblotting assay proved to be a useful tool for determining antigenic markers of recent and chronic phases of
Toxoplasma gondii
infection.
Molecular Confirmation of Human Immunodeficiency Virus (HIV) Type 2 in HIV-Seropositive Subjects in South India
Authors: Kannangai R. , Ramalingam S. , Prakash K. J. , Abraham O. C. , George R. , Castillo R. C. , Schwartz D. H. , Jesudason M. V. , Sridharan G. 
Published: July 2002

ABSTRACT
Nested PCRs for human immunodeficiency virus type 1 (HIV-1) and HIV-2 were compared with immunoblot test results. Twelve of 13 immunoblot-positive HIV-2 samples were positive by PCR. There were five INNO-LIA (Innogenetics, Zwijnaarde, Belgium) and/or HIVBLOT 2.2 (Genelabs, Singapore) samples that tested positive for dual infection. HIV-1 PCR was positive in all samples, while HIV-2 PCR was positive in two and RIBA (Chiron Corporation, San Diego, Calif.) was positive for HIV-2 in three samples. Thus the prevalence of HIV-2 is accurately estimated by the use of immunoblotting, but that of HIV-1 and -2 dual infection may be overestimated.
Comparison of Specific Serological Assays for Diagnosing Human Herpesvirus 6 Infection after Liver Transplantation
Authors: Yoshikawa Tetsushi , Black Jodi B. , Ihira Masaru , Suzuki Kyoko , Suga Sadao , Iida Keiji , Saito Yumiko , Asonuma Katsuhiro , Tanaka Koichi , Asano Yoshizo 
Published: July 2002

ABSTRACT
Cross-reactivity between human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7) antibodies and the reliability of specific serological assays were analyzed for 12 patients with concurrent HHV-6 and HHV-7 antibody responses after transplantation with a liver from a living relative by using an immunofluorescence assay (IFA). A neutralizing antibody titer assay (NT) and an immunoblot assay (IB) designed to detect immunoglobulin M (IgM) antibody to the HHV-6 immunodominant 101-kDa protein were compared in the diagnosis of an active HHV-6 infection. A total of 9 of 12 patients demonstrated concurrent HHV-6 and HHV-7 antibody responses, including increased IgG titers and/or the presence of IgM by IFA, and were thus analyzed for cross-reactive antibody to heterologous virus. The average percentages of residual antibody to HHV-6 and HHV-7 after absorption with HHV-6 antigen were 32.6% (range, 6 to 50%) and 55.6% (range, 35 to 100%), respectively. All 12 patients were subsequently analyzed for HHV-6 antibody by using IB and NT. IB detected IgM antibody to the 101-kDa protein in 75% (9 of 12) of the recipients. A significant rise in the NT antibody titer was detected in the same nine samples. However, HHV-6 DNA was detected by PCR in only five of nine plasma samples collected from recipients with a specific serologic response against HHV-6.
Effect of Immunization with Recombinant OspA on Serologic Tests for Lyme Borreliosis
Authors: Fawcett Paul T. , Rose Carlos D. , Budd Sandra M. , Gibney Kathleen M. 
Published: July 2002

ABSTRACT

This study evaluated the effects of vaccination with OspA on the use of serologic tests as aids in the diagnosis of Lyme borreliosis. Sera from control and OspA-immunized mice and from OspA-immunized human volunteers were tested for serologic reactivity to
Borrelia burgdorferi
. Testing was performed with samples obtained prior to administration of vaccine and at 30 days following administration of an initial and a second dose of OspA vaccine. The assays used to assess serologic reactivity included an in-house-developed enzyme-linked immunosorbent assay (ELISA), an in-house-developed Western blot assay, two commercial Western blot tests, and a commercially available dot blot assay. Data obtained from this study demonstrate that immunization with the OspA vaccine will cause ELISA to yield positive results (as reported previously) for the majority of vaccine recipients. Results obtained from Western blot analysis indicate that vaccination with recombinant OspA induces production of antibodies which bind to several different borrelial proteins. The degree of reactivity detected by Western blotting varied greatly between the three assays used. The in-house assay showed the least reactivity, while one commercial Western blot test actually yielded positive test results for infection with
B. burgdorferi
. The usefulness of all three Western blot assays for the diagnosis of potential infection in a vaccine recipient is severely limited by the extensive reactivity caused by vaccination alone. Antibodies produced in response to OspA vaccination did not significantly affect the performance of the dot blot test; thus, it could provide a reliable means to test for infection with
B. burgdorferi
in OspA vaccine recipients.
Optimization of a Human Papillomavirus-Specific Enzyme-Linked Immunosorbent Assay
Authors: Karem Kevin L. , Poon Alysia C. , Bierl Cynthia , Nisenbaum Rosane , Unger Elizabeth 
Published: July 2002

ABSTRACT
A strategy was developed for the control, standardization, and critical evaluation of an enzyme-linked immunosorbent assay (ELISA) for the detection of human papillomavirus-specific immunoglobulin G in human sera. Control human sera, polyclonal animal sera, and monoclonal antibodies were used to establish optimal assay parameters, including antigen coating, serum dilutions, and criteria for daily reproducibility, monitoring, and rejection of assays. Three evaluation techniques were used in parallel to define an optimal cutoff absorbance value that yields greater than 93% sensitivity and 98.5% specificity in the assay's ability to discriminate positive and negative control sera. This strategy provides an optimal method by which to determine cutoff absorbance values for ELISA.
Comparison of Immunoglobulin G Subclass Profiles Induced by Measles Virus in Vaccinated and Naturally Infected Individuals
Authors: Isa María Beatríz , Martínez Laura , Giordano Miguel , Passeggi Carlos , de Wolff María Cristina , Nates Silvia 
Published: July 2002

ABSTRACTA total of 258 human sera positive for measles antibodies were divided into four different groups: group 1 contained 54 sera from children after natural measles infection (immunoglobulin M [IgM] positive, early infection phase), group 2 contained 28 sera from children after measles vaccination (IgM positive, early infection phase), group 3 contained 100 sera from healthy adults (natural long-lasting immunity), and group 4 contained 76 sera from healthy children (postvaccinal long-lasting immunity). In the early phase of infection, the percent distributions of measles virus-specific IgG isotypes were similar between natural and postvaccinal immune responses. IgG1 and IgG4 were the dominant isotypes, with mean levels of detection of 100% (natural infection) and 100% (postvaccinal) for IgG1 and 96% (natural infection) and 92% (postvaccinal) for IgG4. In comparison, the IgG4 geometric mean titer (GMT) in the early phase of natural infection was significantly higher than the IgG4 GMT detected in the postvaccinal immune response (80 versus 13; 95% confidence interval). In the memory phase, IgG2 and IgG3 responses decreased significantly in both natural infection and postvaccinal groups, while IgG1 levels were maintained. In contrast, the IgG4 postvaccinal immune response decreased strongly in the memory phase, whereas IgG4 natural long-lasting immunity remained unchanged (9 versus 86%;P< 0.05). The results obtained suggest that IgG4 isotype could be used in the early phase of infection as a quantitative marker and in long-lasting immunity as a qualitative marker to differentiate between natural and postvaccinal immune responses.
Cellular Immune Responses to Recombinant Antigens in Pregnant Women Chronically Infected with
Toxoplasma gondii
Authors: Fatoohi A. F. , Cozon G. J. N. , Greenland T. , Ferrandiz J. , Bienvenu J. , Picot S. , Peyron F. 
Published: July 2002

ABSTRACT

The parasite
Toxoplasma gondii
can infect most mammals and birds, sometimes causing severe pathology. Primary infection during pregnancy can result in abortion or fetal defects. Host immunity, particularly cellular immunity towards antigenic peptides, can control infection, but an efficient vaccine is not yet available. We have evaluated T-cell responses to a crude soluble toxoplasma antigen (ST-Ag) and to five recombinant peptide antigens of cells in whole-blood cultures from 22 pregnant women with preexisting infections and from 7 pregnant negative controls. Cells from all infected patients but from none of the controls responded specifically to ST-Ag by expressing surface CD25 on culture. Responses to the recombinant antigens showed considerable variation between individuals. rGRA1 elicited a response in 16 of the 22 samples (73%), rSAG1 in 13, rGRA7 in 9, rGRA6-CT in 4, and rGRA6-NT in only 1. Most responding cells were CD4
+
. Cells from infected subjects cultured with ST-Ag all released high levels of gamma interferon (IFN-γ) into the culture supernatant (4,343 ± 2,536 pg/ml). Cells from 12 patients released IFN-γ after culture with rGRA1 (130 ± 98 pg/ml), those from 10 patients released it after culture with rSAG1 (183 ± 128 pg/ml), and those from 4 patients released it after culture with rGRA7 (324 ± 374 pg/ml). Intensity of IFN-γ production in response to the latter two recombinant antigens correlated with responses to ST-Ag (
r
= 0.61 and 0.53, respectively;
P
< 0.01). Interleukin-4 was always absent from supernatants of cells stimulated with toxoplasma antigens. The heterogeneity of human responses to individual recombinant toxoplasma antigens should be considered in the design of potential vaccines.
Salmonella
Pathogenicity Island 1-Independent Induction of Apoptosis in Infected Macrophages by
Salmonella enterica
Serotype Typhimurium
Authors: van der Velden Adrianus W. M. , Lindgren Susanne W. , Worley Micah J. , Heffron Fred 
Published: July 2002

ABSTRACT

The enteric pathogen
Salmonella enterica
serotype Typhimurium induces apoptosis in infected macrophages. This process is rapid, specific, and depends on the type III protein secretion system encoded within
Salmonella
pathogenicity island 1 (SPI1). Here, we demonstrate that serotype Typhimurium can activate programmed macrophage cell death independently of SPI1. SPI1 independent induction of apoptosis in infected macrophages is observed as early as 12 to 13 h postinfection, even in the absence of intracellular bacterial replication. Delayed activation of programmed macrophage cell death is not observed with serotype Typhimurium strains mutated in
ompR
or SPI2. Even though SPI2 mutants have a defect in intracellular proliferation, our results indicate that long-term intracellular survival and growth are not required for delayed macrophage killing per se, since
Salmonella
mutants that are severely defective in intracellular growth still induce delayed apoptosis. Inactivation of genes required for either rapid or delayed induction of apoptosis results in a conditional noncytotoxic phenotype, whereas simultaneous inactivation of genes required for both rapid and delayed induction of apoptosis renders serotype Typhimurium noncytotoxic under all conditions tested. Our hypothesis is that differential activation of programmed macrophage cell death by serotype Typhimurium occurs under discrete physiological conditions at distinct locations within an infected host.
Infection of Human Monocyte-Derived Macrophages withChlamydia trachomatisInduces Apoptosis of T Cells: a Potential Mechanism for Persistent Infection
Authors: Jendro Michael C. , Deutsch Tobias , Körber Britta , Köhler Lars , Kuipers Jens G. , Krausse-Opatz Birgit , Westermann Jürgen , Raum Elke , Zeidler Henning 
Published: July 2002

ABSTRACTViruses can escape T-cell surveillance by infecting macrophages and thereby induce apoptosis of noninfected T cells. This ability had not been demonstrated for bacteria. We investigated whether infection of macrophages with the important human pathogenChlamydia trachomatiscan induce T-cell apoptosis. BecauseChlamydia-Mycoplasmacoinfection is a frequent event, the ability ofMycoplasma fermentans-infected macrophages to induce T-cell apoptosis was also studied. Infected macrophages were cocultivated with autologous T cells in different activation states. Propidium iodide-based fluorescence-activated cell sorter analysis demonstrated that macrophages infected with viable chlamydiae induced T-cell death. Apoptosis was identified as the mode of death induction by using a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling assay. Induction of T-cell death was macrophage dependent. Incubation of T cells with infectious chlamydiae in the absence of macrophages did not lead to T-cell apoptosis. UV irradiation of chlamydiae diminished the ability to induce death. T-cell death was induced by a cell-free supernatant of infected macrophages. Not only phytohemagglutinin-preactivated T cells but also non-mitogen-preactivated T cells were susceptible toC. trachomatis-induced apoptosis. In contrast,M. fermentansinfection of macrophages did not induce T-cell death. Coinfection had no additional effect. In summary, intracellular chlamydial infection of macrophages can induce T-cell apoptosis. Apoptosis induction by chlamydiae possibly explains how persistently infected macrophages escape T-cell surveillance and why theChlamydia-specific T-cell response is diminished during persistent chlamydial infection.