bluuebunny/crossref_metadata_2025_split huggingface dataset:
Kinetic Analysis of Bifidobacterial Metabolism Reveals a Minor Role for Succinic Acid in the Regeneration of NAD
+
through Its Growth-Associated Production
Authors: Van der Meulen Roel , Adriany Tom , Verbrugghe Kristof , De Vuyst Luc 
Published: August 2006

ABSTRACT

Several strains belonging to the genus
Bifidobacterium
were tested to determine their abilities to produce succinic acid.
Bifidobacterium longum
strain BB536 and
Bifidobacterium animalis
subsp.
lactis
strain Bb 12 were kinetically analyzed in detail using in vitro fermentations to obtain more insight into the metabolism and production of succinic acid by bifidobacteria. Changes in end product formation in strains of
Bifidobacterium
could be related to the specific rate of sugar consumption. When the specific sugar consumption rate increased, relatively more lactic acid and less acetic acid, formic acid, and ethanol were produced, and vice versa. All
Bifidobacterium
strains tested produced small amounts of succinic acid; the concentrations were not more than a few millimolar. Succinic acid production was found to be associated with growth and stopped when the energy source was depleted. The production of succinic acid contributed to regeneration of a small part of the NAD
+
, in addition to the regeneration through the production of lactic acid and ethanol.
Multiple-Subunit Genes of the Aromatic-Ring-Hydroxylating Dioxygenase Play an Active Role in Biphenyl and Polychlorinated Biphenyl Degradation in
Rhodococcus
sp. Strain RHA1
Authors: Iwasaki Takumi , Miyauchi Keisuke , Masai Eiji , Fukuda Masao 
Published: August 2006

ABSTRACT

A gram-positive strong polychlorinated biphenyl (PCB) degrader,
Rhodococcus
sp. strain RHA1, can degrade PCBs by cometabolism with biphenyl or ethylbenzene. In RHA1, three sets of aromatic-ring-hydroxylating dioxygenase genes are induced by biphenyl. The large and small subunits of their terminal dioxygenase components are encoded by
bphA1
and
bphA2
,
etbA1
and
etbA2
, and
ebdA1
and
ebdA2
, respectively, and the deduced amino acid sequences of
etbA1
and
etbA2
are identical to those of
ebdA1
and
ebdA2
, respectively. In this study, we examined the involvement of the respective subunit genes in biphenyl/PCB degradation by RHA1. Reverse transcription-PCR and two-dimensional polyacrylamide gel electrophoresis analyses indicated the induction of RNA and protein products of
etbA1
and
ebdA1
by biphenyl. Single- and double-disruption mutants of
etbA1
,
ebdA1
, and
bphA1
were constructed by insertional inactivation. The 4-chlorobiphenyl (4-CB) degradation activities of all the mutants were lower than that of RHA1. The results indicated that all of these genes are involved in biphenyl/PCB degradation. Furthermore, we constructed disruption mutants of
ebdA3
and
bphA3
, encoding ferredoxin, and
etbA4
, encoding ferredoxin reductase components. The 4-CB degradation activities of these mutants were also lower than that of RHA1, suggesting that all of these genes play a role in biphenyl/PCB degradation. The substrate preferences of
etbA1A2
/
ebdA1A2
- and
bphA1A2
-encoded dioxygenases for PCB congeners were examined using the corresponding mutants. The results indicated that these dioxygenase isozymes have different substrate preferences and that the
etbA1A2
/
ebdA1A2
-encoded isozyme is more active on highly chlorinated congeners than the
bphA1A2
-encoded one.
Rickettsia felisfrom Cat Fleas: Isolation and Culture in a Tick-Derived Cell Line
Authors: Pornwiroon Walairat , Pourciau Susan S. , Foil Lane D. , Macaluso Kevin R. 
Published: August 2006

ABSTRACTRickettsia felis, the etiologic agent of spotted fever, is maintained in cat fleas by vertical transmission and resembles other tick-borne spotted fever group rickettsiae. In the present study, we utilized anIxodes scapularis-derived tick cell line, ISE6, to achieve isolation and propagation ofR. felis. A cytopathic effect of increased vacuolization was commonly observed inR. felis-infected cells, while lysis of host cells was not evident despite large numbers of rickettsiae. Electron microscopy identified rickettsia-like organisms in ISE6 cells, and sequence analyses of portions of the citrate synthase (gltA), 16S rRNA,Rickettsiagenus-specific 17-kDa antigen, and spotted fever group-specific outer membrane protein A (ompA) genes and, notably,R. felisconjugative plasmids indicate that this cultivatable strain (LSU) wasR. felis. Establishment ofR. felis(LSU) in a tick-derived cell line provides an alternative and promising system for the expansion of studies investigating the interactions betweenR. felisand arthropod hosts.
Peptide aMptD-Mediated Capture PCR for Detection ofMycobacterium aviumsubsp.paratuberculosisin Bulk Milk Samples
Authors: Stratmann Janin , Dohmann Karen , Heinzmann Julia , Gerlach Gerald-F. 
Published: August 2006

ABSTRACTA peptide-mediated capture PCR for the detection ofMycobacterium aviumsubsp.paratuberculosisin bulk milk samples was developed and characterized. Capture of the organism was performed using peptide aMptD, which had been shown to bind to theM. aviumsubsp.paratuberculosisMptD protein (J. Stratmann, B. Strommenger, R. Goethe, K. Dohmann, G. F. Gerlach, K. Stevenson, L. L. Li, Q. Zhang, V. Kapur, and T. J. Bull, Infect. Immun.72:1265-1274, 2004). Consistent expression of the MptD receptor protein and binding of the aMptD ligand were demonstrated by capturing differentMycobacterium aviumsubsp.paratuberculosistype I and type II strains and subsequent PCR analysis using ISMav2-based primers. The analytical sensitivity of the method was determined to be 5 × 102CFU ml−1for artificially contaminated milk. The specificity of aMptD binding was confirmed by culture and competitive capture assays, showing selective enrichment ofM. aviumsubsp.paratuberculosis(at a concentration of 5 × 102CFU ml−1) from samples containing 100- and 1,000-fold excesses of other mycobacterial species, includingM. aviumsubsp.aviumandM. aviumsubsp.hominissuis. The aMptD-mediated capture ofM. aviumsubsp.paratuberculosisusing paramagnetic beads, followed by culture, demonstrated the ability of this approach to capture viable target cells present in artificially contaminated milk. Surface plasmon resonance experiments revealed that the aMptD peptide is a high-affinity ligand with a calculated association rate constant of 9.28 × 103and an association constant of 1.33 × 109. The potential use of the method on untreated raw milk in the field was investigated by testing 423 bulk milk samples obtained from different dairy farms in Germany, 23 of which tested positive. Taken together, the results imply that the peptide-mediated capture PCR might present a suitable test for paratuberculosis screening of dairy herds, as it has an analytical sensitivity sufficient for detection ofM. aviumsubsp.paratuberculosisin bulk milk samples under field conditions, relies on a defined and validated ligand-receptor interaction, and is adaptable to routine diagnostic laboratory automation.
Dominant Microbial Populations in Limestone-Corroding Stream Biofilms, Frasassi Cave System, Italy
Authors: Macalady Jennifer L. , Lyon Ezra H. , Koffman Bess , Albertson Lindsey K. , Meyer Katja , Galdenzi Sandro , Mariani Sandro 
Published: August 2006

ABSTRACT

Waters from an extensive sulfide-rich aquifer emerge in the Frasassi cave system, where they mix with oxygen-rich percolating water and cave air over a large surface area. The actively forming cave complex hosts a microbial community, including conspicuous white biofilms coating surfaces in cave streams, that is isolated from surface sources of C and N. Two distinct biofilm morphologies were observed in the streams over a 4-year period. Bacterial 16S rDNA libraries were constructed from samples of each biofilm type collected from Grotta Sulfurea in 2002. β-, γ-, δ-, and ε-proteobacteria in sulfur-cycling clades accounted for ≥75% of clones in both biofilms. Sulfate-reducing and sulfur-disproportionating δ-proteobacterial sequences in the clone libraries were abundant and diverse (34% of phylotypes). Biofilm samples of both types were later collected at the same location and at an additional sample site in Ramo Sulfureo and examined, using fluorescence in situ hybridization (FISH). The biomass of all six stream biofilms was dominated by filamentous γ-proteobacteria with
Beggiatoa
-like and/or
Thiothrix
-like cells containing abundant sulfur inclusions. The biomass of ε-proteobacteria detected using FISH was consistently small, ranging from 0 to less than 15% of the total biomass. Our results suggest that S cycling within the stream biofilms is an important feature of the cave biogeochemistry. Such cycling represents positive biological feedback to sulfuric acid speleogenesis and related processes that create subsurface porosity in carbonate rocks.
Metabolic Profiles and Genetic Diversity of Denitrifying Communities in Activated Sludge after Addition of Methanol or Ethanol
Authors: Hallin Sara , Throbäck Ingela Noredal , Dicksved Johan , Pell Mikael 
Published: August 2006

ABSTRACT

External carbon sources can enhance denitrification rates and thus improve nitrogen removal in wastewater treatment plants. The effects of adding methanol and ethanol on the genetic and metabolic diversity of denitrifying communities in activated sludge were compared using a pilot-scale plant with two parallel lines. A full-scale plant receiving the same municipal wastewater, but without external carbon source addition, was the reference. Metabolic profiles obtained from potential denitrification rates with 10 electron donors showed that the denitrifying communities altered their preferences for certain compounds after supplementation with methanol or ethanol and that methanol had the greater impact. Clone libraries of
nirK
and
nirS
genes, encoding the two different nitrite reductases in denitrifiers, revealed that methanol also increased the diversity of denitrifiers of the
nirS
type, which indicates that denitrifiers favored by methanol were on the rise in the community. This suggests that there might be a niche differentiation between
nirS
and
nirK
genotypes during activated sludge processes. The composition of
nirS
genotypes also varied greatly among all samples, whereas the
nirK
communities were more stable. The latter was confirmed by denaturing gradient gel electrophoresis of
nirK
communities on all sampling occasions. Our results support earlier hypotheses that the compositions of denitrifier communities change during predenitrification processes when external carbon sources are added, although no severe effect could be observed from an operational point of view.
Cloning and Expression of Two Crystal Protein Genes,
cry30Ba1
and
cry44Aa1
, Obtained from a Highly Mosquitocidal Strain,
Bacillus thuringiensis
subsp.
entomocidus
INA288
Authors: Ito Takeshi , Ikeya Tomonori , Sahara Ken , Bando Hisanori , Asano Shin-ichiro 
Published: August 2006

ABSTRACT

Two novel crystal protein genes,
cry30Ba
and
cry44Aa
, were cloned from
Bacillus thuringiensis
subsp.
entomocidus
INA288 and expressed in an acrystalliferous strain. Cry44Aa crystals were highly toxic to second-instar
Culex pipiens pallens
(50% mortality concentration [LC
50
] = 6 ng/ml) and
Aedes aegypti
(LC
50
= 12 ng/ml); however, Cry30Ba crystals were not toxic.
Molecular Fingerprinting of
Cryptosporidium
Oocysts Isolated during Water Monitoring
Authors: Nichols Rosely A. B. , Campbell Brian M. , Smith Huw V. 
Published: August 2006

ABSTRACT

We developed and validated a PCR-based method for identifying
Cryptosporidium
species and/or genotypes present on oocyst-positive microscope slides. The method involves removing coverslips and oocysts from previously examined slides followed by DNA extraction. We tested four loci, the 18S rRNA gene (N18SDIAG and N18SXIAO), the
Cryptosporidium
oocyst wall protein (COWP) gene (STN-COWP), and the dihydrofolate reductase (
dhfr
) gene (by multiplex allele-specific PCR), for amplifying DNA from low densities of
Cryptosporidium parvum
oocysts experimentally seeded onto microscope slides. The N18SDIAG locus performed consistently better than the other three tested. Purified oocysts from humans infected with
C. felis
,
C. hominis
, and
C. parvum
and commercially purchased
C. muris
were used to determine the sensitivities of three loci (N18SDIAG, STN-COWP, and N18SXIAO) to detect low oocyst densities. The N18SDIAG primers provided the greatest number of positive results, followed by the N18SXIAO primers and then the STN-COWP primers. Some oocyst-positive slides failed to generate a PCR product at any of the loci tested, but the limit of sensitivity is not entirely based on oocyst number. Sixteen of 33 environmental water monitoring
Cryptosporidium
slides tested (oocyst numbers ranging from 1 to 130) contained mixed
Cryptosporidium
species. The species/genotypes most commonly found were
C. muris
or
C. andersoni
,
C. hominis
or
C. parvum
, and
C. meleagridis
or
Cryptosporidium
sp. cervine, ferret, and mouse genotypes. Oocysts on one slide contained
Cryptosporidium
muskrat genotype II DNA.
Peptidoglycan from
Bacillus cereus
Mediates Commensalism with Rhizosphere Bacteria from the
Cytophaga-Flavobacterium
Group
Authors: Peterson Snow Brook , Dunn Anne K. , Klimowicz Amy K. , Handelsman Jo 
Published: August 2006

ABSTRACT

Previous research in our laboratory revealed that the introduction of
Bacillus cereus
UW85 can increase the populations of bacteria from the
Cytophaga-Flavobacterium
(CF) group of the
Bacteroidetes
phylum in the soybean rhizosphere, suggesting that these rhizosphere microorganisms have a beneficial relationship (G. S. Gilbert, J. L. Parke, M. K. Clayton, and J. Handelsman, Ecology 74:840-854, 1993). In the present study, we determined the frequency at which CF bacteria coisolated with
B. cereus
strains from the soybean rhizosphere and the mechanism by which
B. cereus
stimulates the growth of CF rhizosphere strains in root exudate media. In three consecutive years of sampling, CF strains predominated among coisolates obtained with
B. cereus
isolates from field-grown soybean roots. In root exudate media, the presence of
B. cereus
was required for CF coisolate strains to reach high population density. However, rhizosphere isolates from the phylum
Proteobacteria
grew equally well in the presence and absence of
B. cereus
, and the presence of CF coisolates did not affect the growth of
B. cereus
. Peptidoglycan isolated from
B. cereus
cultures stimulated growth of the CF rhizosphere bacterium
Flavobacterium johnsoniae
, although culture supernatant from
B. cereus
grown in root exudate media did not. These results suggest
B. cereus
and CF rhizosphere bacteria have a commensal relationship in which peptidoglycan produced by
B. cereus
stimulates the growth of CF bacteria.
σBActivation under Environmental and Energy Stress Conditions inListeria monocytogenes
Authors: Chaturongakul Soraya , Boor Kathryn J. 
Published: August 2006

ABSTRACTTo measure σBactivation inListeria monocytogenesunder environmental or energy stress conditions, quantitative reverse transcriptase PCR (TaqMan) was used to determine the levels of transcripts for the σB-dependentopuCAandclpCgenes in strains having null mutations in genes encodingregulator ofsigmaBproteins (rsbTandrsbV) and sigma B (sigB) and in theL. monocytogeneswild-type 10403S strain under different stress conditions. The ΔsigB, ΔrsbT, and ΔrsbVstrains previously exhibited increased hemolytic activities compared to the hemolytic activity of the wild-type strain; therefore, transcript levels forhlywere also determined. RsbT, RsbV, and σBwere all required foropuCAexpression during growth under carbon-limiting conditions or following exposure to pH 4.5, salt, ethanol, or the protonophore carbonyl cyanidem-chlorophenylhydrazone (CCCP). Expression ofclpCwas RsbT, RsbV, and σBdependent in the presence of CCCP but not under the other conditions.hlyexpression was not RsbT, RsbV, or σBdependent in the presence of either CCCP or salt.opuCAtranscript levels did not increase in the presence of rapidly lethal stresses (i.e., pH 2.5 or 13 mM cumene hydroperoxide) despite the enhanced survival of the wild type compared with the survival of the mutant strains under these conditions. These findings highlight the importance of complementing phenotypic characterizations with gene expression studies to identify direct and indirect effects of null mutations in regulatory genes, such assigB. Overall, our data show that while σBactivation occurs through a single pathway under both environmental and energy stress conditions, regulation of expression of some stress response and virulence genes in the σBregulon (e.g.,clpC) appears to require networks involving multiple transcriptional regulators.
Examination of breast needle core biopsy specimens performed for screen-detected microcalcification
Authors: Kumaraswamy V , Carder P J 
Published: August 2006

Aims:To establish the number of histological levels necessary for the evaluation of breast needle core biopsy (NCB) specimens taken from areas of mammographic calcification in patients presenting via the UK National Health Service Breast Screening Programme.Methods:Retrospective review of a series of breast NCB specimens initially examined routinely at nine levels. The presence of calcification within the histological sections in each of three sets of levels (levels 1–3, 4–6 and 7–9) and the (cumulative) diagnostic B category that would have pertained after each were assessed.Results:Accurate diagnostic classification was possible after examination of three levels in 89% cases. Examination of a further three levels permitted accurate diagnosis in a further eight cases (total 97% cases). In only three cases were nine levels necessary for accurate classification. In only a single case (1%) was it likely that routine examination of six levels could have led to significant misclassification. In a significant group of patients (18%), nine levels were considered to provide additional useful information, although this information did not alter the diagnosis.Conclusions:NCBs for screen-detected mammographic calcification should be routinely examined at six levels. Further levels may be needed in occasional cases to identify more conclusively the associated pathological abnormality. Further levels may be of particular value when assessing atypical intraductal proliferative epithelial lesions.
Effect of Target Dynamics on Pharmacokinetics of a Novel Therapeutic Antibody against the Epidermal Growth Factor Receptor: Implications for the Mechanisms of Action
Authors: Lammerts van Bueren Jeroen J. , Bleeker Wim K. , Bøgh Henrik O. , Houtkamp Mischa , Schuurman Janine , van de Winkel Jan G.J. , Parren Paul W.H.I. 
Published: August 2006

AbstractThe epidermal growth factor receptor (EGFR) is overexpressed on many solid tumors and represents an attractive target for antibody therapy. Here, we describe the effect of receptor-mediated antibody internalization on the pharmacokinetics and dose-effect relationship of a therapeutic monoclonal antibody (mAb) against EGFR (2F8). This mAb was previously found therapeutically active in mouse tumor models by two dose-dependent mechanisms of action: blockade of ligand binding and induction of antibody-dependent cell-mediated cytotoxicity. In vitro studies showed 2F8 to be rapidly internalized by EGFR-overexpressing cells. In vivo, accelerated 2F8 clearance was observed in cynomolgus monkeys at low doses but not at high doses. This enhanced clearance seemed to be receptor dependent and was included in a pharmacokinetic model designed to explain its nonlinearity. Receptor-mediated clearance was also found to affect in situ antibody concentrations in tumor tissue. Ex vivo analyses of xenograft tumors of 2F8-treated nude mice revealed that relatively high antibody plasma concentrations were required for maximum EGFR saturation in high-EGFR-expressing human A431 tumors, in contrast to lower-EGFR-expressing human xenograft tumors. In summary, receptor-mediated antibody internalization and degradation provides a saturable route of clearance that significantly affects pharmacokinetics, particularly at low antibody doses. EGFR saturation in normal tissues does not predict saturation in tumor tissue as local antibody concentrations in EGFR-overexpressing tumors may be more rapidly reduced by antibody internalization. Consequently, antibody saturation of the receptor may be affected, thereby affecting the local mechanism of action. (Cancer Res 2006; 66(15): 7630-8)
Androgens Induce Prostate Cancer Cell Proliferation through Mammalian Target of Rapamycin Activation and Post-transcriptional Increases in Cyclin D Proteins
Authors: Xu Youyuan , Chen Shao-Yong , Ross Kenneth N. , Balk Steven P. 
Published: August 2006

AbstractAndrogen receptor (AR) plays a central role in prostate cancer, with most tumors responding to androgen deprivation therapies, but the molecular basis for this androgen dependence has not been determined. Androgen [5α-dihydrotestosterone (DHT)] stimulation of LNCaP prostate cancer cells, which have constitutive phosphatidylinositol 3-kinase (PI3K)/Akt pathway activation due to PTEN loss, caused increased expression of cyclin D1, D2, and D3 proteins, retinoblastoma protein hyperphosphorylation, and cell cycle progression. However, cyclin D1 and D2 message levels were unchanged, indicating that the increases in cyclin D proteins were mediated by a post-transcriptional mechanism. This mechanism was identified as mammalian target of rapamycin (mTOR) activation. DHT treatment increased mTOR activity as assessed by phosphorylation of the downstream targets p70 S6 kinase and 4E-BP1, and mTOR inhibition with rapamycin blocked the DHT-stimulated increase in cyclin D proteins. Significantly, DHT stimulation of mTOR was not mediated through activation of the PI3K/Akt or mitogen-activated protein kinase/p90 ribosomal S6 kinase pathways and subsequent tuberous sclerosis complex 2/tuberin inactivation or by suppression of AMP-activated protein kinase. In contrast, mTOR activation by DHT was dependent on AR-stimulated mRNA synthesis. Oligonucleotide microarrays showed that DHT-stimulated rapid increases in multiple genes that regulate nutrient availability, including transporters for amino acids and other organic ions. These results indicate that a critical function of AR in PTEN-deficient prostate cancer cells is to support the pathologic activation of mTOR, possibly by increasing the expression of proteins that enhance nutrient availability and thereby prevent feedback inhibition of mTOR. (Cancer Res 2006; 66(15): 7783-92)