bluuebunny/crossref_metadata_2025_split huggingface dataset:
Modeling and Predicting the Simultaneous Growth ofEscherichia coliO157:H7 and Ground Beef Background Microflora for Various Enrichment Protocols
Authors: Vimont A. , Vernozy-Rozand C. , Montet M. P. , Lazizzera C. , Bavai C. , Delignette-Muller M.-L. 
Published: January 2006

ABSTRACTThe simultaneous growth ofEscherichia coliO157:H7 (O157) and the ground beef background microflora (BM) was described in order to characterize the effects of enrichment factors on the growth of these organisms. The different enrichment factors studied were basal medium (Trypticase soy broth andE. colibroth), the presence of novobiocin in the broth, and the incubation temperature (37°C or 40°C). BM and O157 kinetics were simultaneously fitted by using a competitive growth model. The simple competition between the two microfloras implied that O157 growth stopped as soon as the maximal bacterial density in the BM was reached. The present study shows that the enrichment protocol factors had little impact on the simultaneous growth of BM and O157. The selective factors (i.e., bile salts and novobiocin) and the higher incubation temperature (40°C) did not inhibit BM growth, and incubation at 40°C only slightly improved O157 growth. The results also emphasize that when the level of O157 contamination in ground beef is low, the 6-h enrichment step recommended in the immunomagnetic separation protocol (ISO EN 16654) is not sufficient to detect O157 by screening methods. In this case, prior enrichment for approximately 10 h appears to be the optimal duration for enrichment. However, more experiments must be carried out with ground beef packaged in different ways in order to confirm the results obtained in the present study for non-vacuum- and non-modified-atmosphere-packed ground beef.
Display of α-Amylase on the Surface of
Lactobacillus casei
Cells by Use of the PgsA Anchor Protein, and Production of Lactic Acid from Starch
Authors: Narita Junya , Okano Kenji , Kitao Tomoe , Ishida Saori , Sewaki Tomomitsu , Sung Moon-Hee , Fukuda Hideki , Kondo Akihiko 
Published: January 2006

ABSTRACT

We developed a new cell surface engineering system based on the PgsA anchor protein from
Bacillus subtilis
. In this system, the N terminus of the target protein was fused to the PgsA protein and the resulting fusion protein was expressed on the cell surface. Using this new system, we constructed a novel starch-degrading strain of
Lactobacillus casei
by genetically displaying α-amylase from the
Streptococcus bovis
strain 148 with a FLAG peptide tag (AmyAF). Localization of the PgsA-AmyA-FLAG fusion protein on the cell surface was confirmed by immunofluorescence microscopy and flow cytometric analysis. The lactic acid bacteria which displayed AmyAF showed significantly elevated hydrolytic activity toward soluble starch. By fermentation using AmyAF-displaying
L. casei
cells, 50 g/liter of soluble starch was reduced to 13.7 g/liter, and 21.8 g/liter of lactic acid was produced within about 24 h. The yield in terms of grams of lactic acid produced per gram of carbohydrate utilized was 0.60 g per g of carbohydrate consumed at 24 h. Since AmyA was immobilized on the cells, cells were recovered after fermentation and used repeatedly. During repeated utilization of cells, the lactic acid yield was improved to 0.81 g per g of carbohydrate consumed at 72 h. These results indicate that efficient simultaneous saccharification and fermentation from soluble starch to lactic acid were carried out by recombinant
L. casei
cells with cell surface display of AmyA.
Role of Autochthonous Filamentous Fungi in Bioremediation of a Soil Historically Contaminated with Aromatic Hydrocarbons
Authors: D'Annibale A. , Rosetto F. , Leonardi V. , Federici F. , Petruccioli M. 
Published: January 2006

ABSTRACT

Nine fungal strains isolated from an aged and heavily contaminated soil were identified and screened to assess their degradative potential. Among them,
Allescheriella
sp. strain DABAC 1,
Stachybotrys
sp. strain DABAC 3, and
Phlebia
sp. strain DABAC 9 were selected for remediation trials on the basis of Poly R-478 decolorization associated with lignin-modifying enzyme (LME) production. These autochthonous fungi were tested for the abilities to grow under nonsterile conditions and to degrade various aromatic hydrocarbons in the same contaminated soil. After 30 days, fungal colonization was clearly visible and was confirmed by ergosterol determination. In spite of subalkaline pH conditions and the presence of heavy metals, the autochthonous fungi produced laccase and Mn and lignin peroxidases. No LME activities were detected in control microcosms. All of the isolates led to a marked removal of naphthalene, dichloroaniline isomers,
o
-hydroxybiphenyl, and 1,1′-binaphthalene.
Stachybotrys
sp. strain DABAC 3 was the most effective isolate due to its ability to partially deplete the predominant contaminants 9,10-anthracenedione and 7H-benz[DE]anthracen-7-one. A release of chloride ions was observed in soil treated with either
Allescheriella
sp. strain DABAC 1 or
Stachybotrys
sp. strain DABAC 3, suggesting the occurrence of oxidative dehalogenation. The autochthonous fungi led to a significant decrease in soil toxicity, as assessed by both the
Lepidium sativum
L. germination test and the Collembola mortality test.
Molecular Evidence for Association of
Chlamydiales
Bacteria with Epitheliocystis in Leafy Seadragon (
Phycodurus eques
), Silver Perch (
Bidyanus bidyanus
), and Barramundi (
Lates calcarifer
)
Authors: Meijer Adam , Roholl Paul J. M. , Ossewaarde Jacobus M. , Jones Brian , Nowak Barbara F. 
Published: January 2006

ABSTRACT

Epitheliocystis in leafy seadragon (
Phycodurus eques
), silver perch (
Bidyanus bidyanus
), and barramundi (
Lates calcarifer
), previously associated with chlamydial bacterial infection using ultrastructural analysis, was further investigated by using molecular and immunocytochemical methods. Morphologically, all three species showed epitheliocystis cysts in the gills, and barramundi also showed lymphocystis cysts in the skin. From gill cysts of all three species and from skin cysts of barramundi 16S rRNA gene fragments were amplified by PCR and sequenced, which clustered by phylogenetic analysis together with other chlamydia-like organisms in the order
Chlamydiales
in a lineage separate from the family
Chlamydiaceae
. By using in situ RNA hybridization, 16S rRNA
Chlamydiales
-specific sequences were detected in gill cysts of silver perch and in gill and skin cysts of barramundi. By applying immunocytochemistry, chlamydial antigens (lipopolysaccharide and/or membrane protein) were detected in gill cysts of leafy seadragon and in gill and skin cysts of barramundi, but not in gill cysts of silver perch. In conclusion, this is the first time epitheliocystis agents of leafy seadragon, silver perch and barramundi have been undoubtedly identified as belonging to bacteria of the order
Chlamydiales
by molecular methods. In addition, the results suggested that lymphocystis cysts, known to be caused by iridovirus infection, could be coinfected with the epitheliocystis agent.
Functional Analysis of Unique Class II Insertion Sequence IS
1071
Authors: Sota Masahiro , Yano Hirokazu , Nagata Yuji , Ohtsubo Yoshiyuki , Genka Hiroyuki , Anbutsu Hisashi , Kawasaki Haruhiko , Tsuda Masataka 
Published: January 2006

ABSTRACT

Various xenobiotic-degrading genes on many catabolic plasmids are often flanked by two copies of an insertion sequence, IS
1071
. This 3.2-kb IS element has long (110-bp) terminal inverted repeats (IRs) and a transposase gene that are phylogenetically related to those of the class II transposons. However, the transposition mechanism of IS
1071
has remained unclear. Our study revealed that IS
1071
was only able to transpose at high frequencies in two environmental β-proteobacterial strains,
Comamonas testosteroni
and
Delftia acidovorans
, and not in any of the bacteria examined which belong to the α- and γ-proteobacteria. IS
1071
was found to have the functional features of the class II transposons in that (i) the final product of the IS
1071
transposition was a cointegrate of its donor and target DNA molecules connected by two directly repeated copies of IS
1071
, one at each junction; (ii) a 5-bp duplication of the target sequence was observed at the insertion site; and (iii) a
tnpA
mutation of IS
1071
was efficiently complemented by supplying the wild-type
tnpA
gene in
trans
. Deletion analysis of the IS
1071
IR sequences indicated that nearly the entire region of the IRs was required for its transposition, suggesting that the interaction between the transposase and IRs of IS
1071
might be different from that of the other well-characterized class II transposons.
In Vitro Alginate Polymerization and the Functional Role of Alg8 in Alginate Production by
Pseudomonas aeruginosa
Authors: Remminghorst Uwe , Rehm Bernd H. A. 
Published: January 2006

ABSTRACT

An enzymatic in vitro alginate polymerization assay was developed by using
14
C-labeled GDP-mannuronic acid as a substrate and subcellular fractions of alginate overproducing
Pseudomonas aeruginosa
FRD1 as a polymerase source. The highest specific alginate polymerase activity was detected in the envelope fraction, suggesting that cytoplasmic and outer membrane proteins constitute the functional alginate polymerase complex. Accordingly, no alginate polymerase activity was detected using cytoplasmic membrane or outer membrane proteins, respectively. To determine the requirement of Alg8, which has been proposed as catalytic subunit of alginate polymerase, nonpolar isogenic
alg8
knockout mutants of alginate-overproducing
P. aeruginosa
FRD1 and
P. aeruginosa
PDO300 were constructed, respectively. These mutants were deficient in alginate biosynthesis, and alginate production was restored by introducing only the
alg8
gene. Surprisingly, this resulted in significant alginate overproduction of the complemented
P. aeruginosa
Δalg8 mutants compared to nonmutated strains, suggesting that Alg8 is the bottleneck in alginate biosynthesis.
1
H-NMR analysis of alginate isolated from these complemented mutants showed that the degree of acetylation increased from 4.7 to 9.3% and the guluronic acid content was reduced from 38 to 19%. Protein topology prediction indicated that Alg8 is a membrane protein. Fusion protein analysis provided evidence that Alg8 is located in the cytoplasmic membrane with a periplasmic C terminus. Subcellular fractionation suggested that the highest specific PhoA activity of Alg8-PhoA is present in the cytoplasmic membrane. A structural model of Alg8 based on the structure of SpsA from
Bacillus subtilis
was developed.
Plasmid-Mediated High-Level Gentamicin Resistance among Enteric Bacteria Isolated from Pet Turtles in Louisiana
Authors: Díaz María Alejandra , Cooper Richard Kent , Cloeckaert Axel , Siebeling Ronald John 
Published: January 2006

ABSTRACT

The sale of small turtles is banned by the Food and Drug Administration from the U.S. market due to concerns about their excretion of
Salmonella
spp. To produce a safe pet for the export market, the Louisiana pet turtle industry uses gentamicin sulfate baths (1,000 μg/ml) to eradicate
Salmonella
spp. from turtle eggs. In 1999, we analyzed bacterial samples recovered from turtle farms and found that strains of
Salmonella enterica
subsp.
arizonae
and other bacteria, such as
Enterobacter cloacae
,
Citrobacter freundii
,
Pseudomonas aeruginosa
, and
Stenotrophomonas maltophilia
, were resistant to high concentrations of gentamicin (>2,000 μg/ml) and to other aminoglycosides. The goal of this study was to identify the gene(s) which contributes to the high-level gentamicin resistance phenotype observed in bacteria from environmental samples with turtle farming activity, particularly the salmonellae, and to estimate the incidence of such genes in these bacteria. R plasmids from gentamicin-resistant strains were transferred by conjugation and transformation to naive
Escherichia coli
cells. Cloning and sequencing of the gentamicin resistance determinants on these plasmids revealed the presence of the aminoglycoside acetyltransferase genes
aac(3)-IIa
and
aac(3)-VIa
; the latter was present as a gene cassette of a class 1 integron. Multiplex PCR assays showed that every gentamicin-resistant isolate carried one of these acetyltransferase genes. Pulsed-field gel electrophoresis and restriction enzyme digestion analysis of R plasmids carrying these genes revealed different restriction profiles and sizes, indicating a dissemination of the gentamicin resistance genes through mobile molecular elements. The data presented highlight the need to develop an alternate method for the eradication of
Salmonella
spp. from turtle eggs.
Inhibition of
Listeria monocytogenes
by Food-Borne Yeasts
Authors: Goerges Stefanie , Aigner Ulrike , Silakowski Barbara , Scherer Siegfried 
Published: January 2006

ABSTRACT

Many bacteria are known to inhibit food pathogens, such as
Listeria monocytogenes
, by secreting a variety of bactericidal and bacteriostatic substances. In sharp contrast, it is unknown whether yeast has an inhibitory potential for the growth of pathogenic bacteria in food. A total of 404 yeasts were screened for inhibitory activity against five
Listeria monocytogenes
strains. Three hundred and four of these yeasts were isolated from smear-ripened cheeses. Most of the yeasts were identified by Fourier transform infrared spectroscopy. Using an agar-membrane screening assay, a fraction of approximately 4% of the 304 red smear cheese isolates clearly inhibited growth of
L. monocytogenes
. Furthermore, 14 out of these 304 cheese yeasts were cocultivated with
L. monocytogenes
WSLC 1364 on solid medium to test the antilisterial activity of yeast in direct cell contact with
Listeria
. All yeasts inhibited
L. monocytogenes
to a low degree, which is most probably due to competition for nutrients. However, one
Candida intermedia
strain was able to reduce the listerial cell count by 4 log units. Another four yeasts, assigned to
C. intermedia
(three strains) and
Kluyveromyces marxianus
(one strain), repressed growth of
L. monocytogenes
by 3 log units. Inhibition of
L. monocytogenes
was clearly pronounced in the cocultivation assay, which simulates the conditions and contamination rates present on smear cheese surfaces. We found no evidence that the unknown inhibitory molecule is able to diffuse through soft agar.
Cometabolism of Citrate and Glucose by
Enterococcus faecium
FAIR-E 198 in the Absence of Cellular Growth
Authors: Vaningelgem Frederik , Ghijsels Veerle , Tsakalidou Effie , De Vuyst Luc 
Published: January 2006

ABSTRACT

Citrate metabolism by
Enterococcus faecium
FAIR-E 198, an isolate from Greek Feta cheese, was studied in modified MRS (mMRS) medium under different pH conditions and glucose and citrate concentrations. In the absence of glucose, this strain was able to metabolize citrate in a pH range from constant pH 5.0 to 7.0. At a constant pH 8.0, no citrate was metabolized, although growth took place. The main end products of citrate metabolism were acetate, formate, acetoin, and carbon dioxide, whereas ethanol and diacetyl were present in smaller amounts. In the presence of glucose, citrate was cometabolized, but it did not contribute to growth. Also, more acetate and less acetoin were formed compared to growth in mMRS medium and in the absence of glucose. Most of the citrate was consumed during the stationary phase, indicating that energy generated by citrate metabolism was used for maintenance. Experiments with cell-free fermented mMRS medium indicated that
E. faecium
FAIR-E 198 was able to metabolize another energy source present in the medium.
In Vivo Random Mutagenesis of
Bacillus subtilis
by Use of TnYLB-1, a
mariner
-Based Transposon
Authors: Le Breton Yoann , Mohapatra Nrusingh Prasad , Haldenwang W. G. 
Published: January 2006

ABSTRACT

This report describes the construction and characterization of a
mariner
-based transposon system designed to be used in
Bacillus subtilis
, but potentially applicable to other gram-positive bacteria. Two pUC19-derived plasmids were created that contain the
mariner
-
Himar1
transposase gene, modified for expression in
B. subtilis
, under the control of either σ
A
- or σ
B
-dependent promoters. Both plasmids also contain a transposable element (TnYLB-1) consisting of a Kan
r
cassette bracketed by the
Himar1
-recognized inverse terminal repeats, as well as the temperature-sensitive replicon and Erm
r
gene of pE194ts. TnYLB-1 transposes into the
B. subtilis
chromosome with high frequency (10
−2
) from either plasmid. Southern hybridization analyses of 15 transposants and sequence analyses of the insertion sites of 10 of these are consistent with random transposition, requiring only a “TA” dinucleotide as the essential target in the recipient DNA. Two hundred transposants screened for sporulation proficiency and auxotrophy yielded five Spo
−
clones, three with insertions in known sporulation genes (
kinA
,
spoVT
, and
yqfD
) and two in genes (
ybaN
and
yubB
) with unknown functions. Two auxotrophic mutants were identified among the 200 transposants, one with an insertion in
lysA
and another in a gene (
yjzB
) whose function is unknown.
Construction of Improved Temperature-Sensitive and Mobilizable Vectors and Their Use for Constructing Mutations in the Adhesin-Encoding
acm
Gene of Poorly Transformable Clinical
Enterococcus faecium
Strains
Authors: Nallapareddy Sreedhar R. , Singh Kavindra V. , Murray Barbara E. 
Published: January 2006

ABSTRACT

Inactivation by allelic exchange in clinical isolates of the emerging nosocomial pathogen
Enterococcus faecium
has been hindered by lack of efficient tools, and, in this study, transformation of clinical isolates was found to be particularly problematic. For this reason, a vector for allelic replacement (pTEX5500ts) was constructed that includes (i) the pWV01-based gram-positive
repAts
replication region, which is known to confer a high degree of temperature intolerance, (ii)
Escherichia coli oriR
from pUC18, (iii) two extended multiple-cloning sites located upstream and downstream of one of the marker genes for efficient cloning of flanking regions for double-crossover mutagenesis, (iv) transcriptional terminator sites to terminate undesired readthrough, and (v) a synthetic extended promoter region containing the
cat
gene for allelic exchange and a high-level gentamicin resistance gene,
aph
(
2
′′)
-Id
, to distinguish double-crossover recombination, both of which are functional in gram-positive and gram-negative backgrounds. To demonstrate the functionality of this vector, the vector was used to construct an
acm
(encoding an adhesin to collagen from
E. faecium
) deletion mutant of a poorly transformable multidrug-resistant
E. faecium
endocarditis isolate, TX0082. The
acm
-deleted strain, TX6051 (TX0082Δ
acm
), was shown to lack Acm on its surface, which resulted in the abolishment of the collagen adherence phenotype observed in TX0082. A mobilizable derivative (pTEX5501ts) that contains
oriT
of Tn
916
to facilitate conjugative transfer from the transformable
E. faecalis
strain JH2Sm::Tn
916
to
E. faecium
was also constructed. Using this vector, the
acm
gene of a nonelectroporable
E. faecium
wound isolate was successfully interrupted. Thus, pTEX5500ts and its mobilizable derivative demonstrated their roles as important tools by helping to create the first reported allelic replacement in
E. faecium
; the constructed this
acm
deletion mutant will be useful for assessing the role of
acm
in
E. faecium
pathogenesis using animal models.
Mercury Analysis of Acid- and Alkaline-Reduced Biological Samples: Identification of
meta
-Cinnabar as the Major Biotransformed Compound in Algae
Authors: Kelly David , Budd Kenneth , Lefebvre Daniel D. 
Published: January 2006

ABSTRACT

The biotransformation of Hg
II
in pH-controlled and aerated algal cultures was investigated. Previous researchers have observed losses in Hg detection in vitro with the addition of cysteine under acid reduction conditions in the presence of SnCl
2
. They proposed that this was the effect of Hg-thiol complexing. The present study found that cysteine-Hg, protein and nonprotein thiol chelates, and nucleoside chelates of Hg were all fully detectable under acid reduction conditions without previous digestion. Furthermore, organic (R-Hg) mercury compounds could not be detected under either the acid or alkaline reduction conditions, and only β-HgS was detected under alkaline and not under acid SnCl
2
reduction conditions. The blue-green alga
Limnothrix planctonica
biotransformed the bulk of Hg
II
applied as HgCl
2
into a form with the analytical properties of β-HgS. Similar results were obtained for the eukaryotic alga
Selenastrum minutum
. No evidence for the synthesis of organomercurials such as CH
3
Hg
+
was obtained from analysis of either airstream or biomass samples under the aerobic conditions of the study. An analytical procedure that involved both acid and alkaline reduction was developed. It provides the first selective method for the determination of β-HgS in biological samples. Under aerobic conditions, Hg
II
is biotransformed mainly into β-HgS (
meta
-cinnabar), and this occurs in both prokaryotic and eukaryotic algae. This has important implications with respect to identification of mercury species and cycling in aquatic habitats.
Cloning and Expression of a Xylitol-4-Dehydrogenase Gene from
Pantoea ananatis
Authors: Aarnikunnas J. S. , Pihlajaniemi A. , Palva A. , Leisola M. , Nyyssölä A. 
Published: January 2006

ABSTRACT

The
Pantoea ananatis
ATCC 43072 mutant strain is capable of growing with xylitol as the sole carbon source. The xylitol-4-dehydrogenase (XDH) catalyzing the oxidation of xylitol to
l
-xylulose was isolated from the cell extract of this strain. The N-terminal amino acid sequence of the purified protein was determined, and an oligonucleotide deduced from this peptide sequence was used to isolate the xylitol-4-dehydrogenase gene (
xdh
) from a
P. ananatis
gene library. Nucleotide sequence analysis revealed an open reading frame of 795 bp, encoding the xylitol-4-dehydrogenase, followed by a 5′ region of another open reading frame encoding an unknown protein. Results from a Northern analysis of total RNA isolated from
P. ananatis
ATCC 43072 suggested that
xdh
is transcribed as part of a polycistronic mRNA. Reverse transcription-PCR analysis of the transcript confirmed the operon structure and suggested that
xdh
was the first gene of the operon. Homology searches revealed that the predicted amino acid sequence of the
P. ananatis
XDH shared significant identity (38 to 51%) with members of the short-chain dehydrogenase/reductase family. The
P. ananatis xdh
gene was successfully overexpressed in
Escherichia coli
, XDH was purified to homogeneity, and some of its enzymatic properties were determined. The enzyme had a preference for NAD
+
as the cosubstrate, and in contrast to previous reports, the enzyme also showed a side activity for the
d
-form of xylulose. Xylitol was converted to
l
-xylulose with a high yield (>80%) by the resting recombinant cells, and the
l
-xylulose was secreted into the medium. No evidence of
d
-xylulose being synthesized by the recombinant cells was found.