bluuebunny/crossref_metadata_2025_split huggingface dataset:
Documenting the Epidemiologic Patterns of Polyomaviruses in Human Populations by Studying Their Presence in Urban Sewage
Authors: Bofill-Mas Sílvia , Pina Sonia , Girones Rosina 
Published: April 2009

ABSTRACT

This is the first description, to our knowledge, of the distribution of human polyomavirus and simian virus 40 (SV40) in urban sewage. Using a nested-PCR procedure, we report the detection of human polyomaviruses JC virus (JCV) and BK virus (BKV) but not SV40 in a high percentage of urban sewage samples obtained from widely divergent geographical areas in Europe and Africa. For a total of 28 samples analyzed, JCV was detected in 26, BKV was detected in 22, and none was positive for SV40. All geographical areas showed a high prevalence of these viruses with mean estimated values of JC viral particles per ml on the order of 10
3
in Barcelona (Spain) and Nancy (France) and 10
2
in Pretoria (South Africa) and Umeå (Sweden) and mean values of BK viral particles on the order of 10
2
in Pretoria and Barcelona and 10
1
in Nancy and Umeå. This compares with estimated mean values of 10
2
to 10
3
for human adenovirus that was evaluated as a control. Nucleotide sequence analysis of the amplified DNA from some of the samples is also presented and represents the sequence of the most abundant JC and BK viral strains in these samples. The nucleotide sequence of the JCV detected was also analyzed in a phylogenetic study and for genomic characterization in the regulatory region. This study has shown that human polyomaviruses are spread in high concentrations in the sewage of different geographical areas and are present in contaminated environments. The frequency and concentration of JCV detected in the environment and the absence of described animal hosts suggest that JCV may be useful as a marker for fecal pollution of anthropogenic origin. The results also support the idea previously described that the strains of JCV are closely related to the ethnic origin of the population studied. The procedure applied should also be useful in future studies of population patterns of viral excretion and as a tool in epidemiological studies for the detection of changes in the prevalence of specific viral pathogens.
Comparative Study of Pressure- and Nutrient-Induced Germination of
Bacillus subtilis
Spores
Authors: Wuytack Elke Y. , Soons Johan , Poschet Filip , Michiels Chris W. 
Published: April 2009

ABSTRACT

Germination experiments with specific germination mutants of
Bacillus subtilis
, including a newly isolated mutant affected in pressure-induced germination, suggest that a pressure of 100 MPa triggers the germination cascades that are induced by the nutrient germinant alanine (Ala) and by a mixture of asparagine, glucose, fructose, and potassium ions (AGFK), by activating the receptors for alanine and asparagine, GerA and GerB, respectively. As opposed to germination at 100 MPa, germination at 600 MPa apparently shortcuts at least part of the Ala- and AGFK-induced germination pathways. Inhibitors of nutrient-induced germination (HgCl
2
and
N
α-P-tosyl-
l
-arginine methyl ester) also inhibit pressure-induced germination at 600 MPa, suggesting that germination at 600 MPa involves activation of a true physiological germination pathway and is therefore not merely a physico-chemical process in which water is forced into the spore protoplast.
A Pilot Study of Bacteriological Population Changes through Potable Water Treatment and Distribution
Authors: Norton Cheryl D. , LeChevallier Mark W. 
Published: April 2009

ABSTRACT
This pilot study compares the compositions of bacterial biofilms in pipe networks supplied with water containing either high levels of biodegradable organic matter (BOM) or low levels of BOM (conventionally or biologically treated, respectively). The Microbial Identification System for fatty acid analysis was utilized in this study to identify a large number of organisms (>1,400) to determine population changes in both conventionally and biologically treated water and biofilms. Data generated during this study indicated that suspended bacteria have little impact on biofilms, and despite treatment (conventional or biological), suspended microbial populations were similar following disinfection. Prechlorination with free chlorine resulted not only in reduced plate count values but also in a dramatic shift in the composition of the bacterial population to predominately gram-positive bacteria. Chlorination of biologically treated water produced the same shifts toward gram-positive bacteria. Removal of assimilable organic carbon by the biologically active filters slowed the rate of biofilm accumulation, but biofilm levels were similar to those found in conventionally treated water within several weeks. Iron pipes stimulated the rate of biofilm development, and bacterial levels on disinfected iron pipes exceeded those for chlorinated polyvinyl chloride pipes. The study showed that the iron pipe surface dramatically influenced the composition, activity, and disinfection resistance of biofilm bacteria.
Carbon Catabolite Repression in
Lactobacillus pentosus
: Analysis of the
ccpA
Region
Authors: Mahr Kerstin , Hillen Wolfgang , Titgemeyer Fritz 
Published: April 2009

ABSTRACT

The catabolite control protein CcpA is a central regulator in low-G+C-content gram-positive bacteria. It confers carbon catabolite repression to numerous genes required for carbon utilization. It also operates as a transcriptional activator of genes involved in diverse phenomena, such as glycolysis and ammonium fixation. We have cloned the
ccpA
region of
Lactobacillus pentosus. ccpA
encodes a protein of 336 amino acids exhibiting similarity to CcpA proteins of other bacteria and to proteins of the LacI/GalR family of transcriptional regulators. Upstream of
ccpA
was found an open reading frame with similarity to the
pepQ
gene, encoding a prolidase. Primer extension experiments revealed two start sites of transcription for
ccpA
. In wild-type cells grown on glucose, mRNA synthesis occurred only from the promoter proximal to
ccpA
. In a
ccpA
mutant strain, both promoters were used, with increased transcription from the distant promoter, which overlaps a presumptive CcpA binding site called
cre
(for catabolite responsive element). This suggests that expression of
ccpA
is autoregulated. Determination of the expression levels of CcpA in cells grown on repressing and nonrepressing carbon sources revealed that the amounts of CcpA produced did not change significantly, leading to the conclusion that the arrangement of two promoters may ensure constant expression of
ccpA
under various environmental conditions. A comparison of the genetic structures of
ccpA
regions revealed that lactic acid bacteria possess the gene order
pepQ-ccpA
-variable while the genetic structure in bacilli and
Staphylococcus xylosus
is
aroA-ccpA
-variable-
acuC
.
In Situ Hybridization of
Prochlorococcus
and
Synechococcus
(Marine Cyanobacteria) spp. with rRNA-Targeted Peptide Nucleic Acid Probes
Authors: Worden Alexandra Z. , Chisholm Sallie W. , Binder Brian J. 
Published: April 2009

ABSTRACT

A simple method for whole-cell hybridization using fluorescently labeled rRNA-targeted peptide nucleic acid (PNA) probes was developed for use in marine cyanobacterial picoplankton. In contrast to established protocols, this method is capable of detecting rRNA in
Prochlorococcus
, the most abundant unicellular marine cyanobacterium. Because the method avoids the use of alcohol fixation, the chlorophyll content of
Prochlorococcus
cells is preserved, facilitating the identification of these cells in natural samples. PNA probe-conferred fluorescence was measured flow cytometrically and was always significantly higher than that of the negative control probe, with positive/negative ratio varying between 4 and 10, depending on strain and culture growth conditions.
Prochlorococcus
cells from open ocean samples were detectable with this method. RNase treatment reduced probe-conferred fluorescence to background levels, demonstrating that this signal was in fact related to the presence of rRNA. In another marine cyanobacterium,
Synechococcus
, in which both PNA and oligonucleotide probes can be used in whole-cell hybridizations, the magnitude of fluorescence from the former was fivefold higher than that from the latter, although the positive/negative ratio was comparable for both probes. In
Synechococcus
cells growing at a range of growth rates (and thus having different rRNA concentrations per cell), the PNA- and oligonucleotide-derived signals were highly correlated (
r
= 0.99). The chemical nature of PNA, the sensitivity of PNA-RNA binding to single-base-pair mismatches, and the preservation of cellular integrity by this method suggest that it may be useful for phylogenetic probing of whole cells in the natural environment.
Isolation and Expression in
Escherichia coli
of
cslA
and
cslB
, Genes Coding for the Chondroitin Sulfate-Degrading Enzymes Chondroitinase AC and Chondroitinase B, Respectively, from
Flavobacterium heparinum
Authors: Tkalec Ana Lydia , Fink Dominique , Blain Françoise , Zhang-Sun Guiyi , Laliberte Maryse , Bennett D. Clark , Gu Kangfu , Zimmermann Joseph J. F. , Su Hongsheng 
Published: April 2009

ABSTRACT

In medium supplemented with chondroitin sulfate,
Flavobacterium heparinum
synthesizes and exports two chondroitinases, chondroitinase AC (chondroitin AC lyase; EC
4.2.2.5
) and chondroitinase B (chondroitin B lyase; no EC number), into its periplasmic space. Chondroitinase AC preferentially depolymerizes chondroitin sulfates A and C, whereas chondroitinase B degrades only dermatan sulfate (chondroitin sulfate B). The genes coding for both enzymes were isolated from
F. heparinum
and designated
cslA
(chondroitinase AC) and
cslB
(chondroitinase B). They were found to be separated by 5.5 kb on the chromosome of
F. heparinum
, transcribed in the same orientation, but not linked to any of the heparinase genes. In addition, the synthesis of both enzymes appeared to be coregulated. The
cslA
and
cslB
DNA sequences revealed open reading frames of 2,103 and 1,521 bp coding for peptides of 700 and 506 amino acid residues, respectively. Chondroitinase AC has a signal sequence of 22 residues, while chondroitinase B is composed of 25 residues. The mature forms of chondroitinases AC and B are comprised of 678 and 481 amino acid residues and have calculated molecular masses of 77,169 and 53,563 Da, respectively. Truncated
cslA
and
cslB
genes have been used to produce active, mature chondroitinases in the cytoplasm of
Escherichia coli
. Partially purified recombinant chondroitinases AC and B exhibit specific activities similar to those of chondroitinases AC and B from
F. heparinum.
An Explosive Antisense RNA Strategy for Inhibition of a Lactococcal Bacteriophage
Authors: Walker Shirley A. , Klaenhammer Todd R. 
Published: April 2009

ABSTRACT

The coding regions of six putative open reading frames (ORFs) identified near the phage φ31 late promoter and the right cohesive end (
cos
) of lactococcal bacteriophage φ31 were used to develop antisense constructs to inhibit the proliferation of phage φ31. Two middle-expressed ORFs (ORF 1 and ORF 2) and four late-expressed ORFs (ORF 3 through ORF 6) were cloned individually between the strong
Lactobacillus
P6 promoter and the T7 terminator (T
T7
) to yield a series of antisense RNA transcripts. When expressed on a high-copy-number vector from a strong promoter, the constructs had no effect on the efficiency of plaquing (EOP) or the plaque size of phage φ31. To increase the ratio of antisense RNA to the targeted sense mRNA appearing during a phage infection, the antisense cassettes containing the late-expressed ORFs (ORF 3 through ORF 6) were subcloned to pTRK360, a low-copy-number vector containing the phage φ31 origin of replication,
ori31. ori31
allows for explosive amplification of the low-copy-number vector upon phage infection, thereby increasing levels of antisense RNA transcripts later in the lytic cycle. In addition, the presence of
ori31
also lowers the burst size of phage φ31 fourfold, resulting in fewer sense, target mRNAs being expressed from the phage genome. The combination of
ori31
and P6::anti-ORF 4H::T
T7
resulted in a threefold decrease in the EOP of phage φ31 (EOP = 0.11 ± 0.03 [mean ± standard deviation]) compared to the presence of
ori31
alone (EOP = 0.36). One-step growth curves showed that expression of anti-ORF 4H RNA decreased the percentage of successful centers of infection (75 to 80% for
ori31
compared to 35 to 45% for
ori31
plus anti-ORF 4H), with no further reduction in burst size. Growth curves performed in the presence of varying levels of phage φ31 showed that
ori31
plus anti-ORF 4H offered significant protection to
Lactococcus lactis
, even at multiplicities of infection of 0.01 and 0.1. These results illustrate a successful application of an antisense strategy to inhibit phage replication in the wake of recent unsuccessful reports.
Lack of Host Specialization inAspergillus flavus
Authors: St. Leger Raymond J. , Screen Steven E. , Shams-Pirzadeh Bijan 
Published: April 2009

ABSTRACTAspergillusspp. cause disease in a broad range of organisms, but it is unknown if strains are specialized for particular hosts. We evaluated isolates ofAspergillus flavus,Aspergillus fumigatus, andAspergillus nidulansfor their ability to infect bean leaves, corn kernels, and insects (Galleria mellonella). Strains ofA. flavusdid not affect nonwounded bean leaves, corn kernels, or insects at 22°C, but they killed insects following hemocoelic challenge and caused symptoms ranging from moderate to severe in corn kernels and bean leaves injured during inoculation. The pectinase P2c, implicated in aggressive colonization of cotton bolls, is produced by mostA. flavusisolates, but its absence did not prevent colonization of bean leaves. Proteases have been implicated in colonization of animal hosts. AllA. flavusstrains produced very similar patterns of protease isozymes when cultured on horse lung polymers. Quantitative differences in protease levels did not correlate with the ability to colonize insects. In contrast toA. flavus, strains ofA. nidulansandA. fumigatuscould not invade living insect or plant tissues or resist digestion by insect hemocytes. Our results indicate thatA. flavushas parasitic attributes that are lacking inA. fumigatusandA. nidulansbut that individual strains ofA. flavusare not specialized to particular hosts.
Rhizosphere Microbial Community Structure in Relation to Root Location and Plant Iron Nutritional Status
Authors: Yang Ching-Hong , Crowley David E. 
Published: April 2009

ABSTRACTRoot exudate composition and quantity vary in relation to plant nutritional status, but the impact of the differences on rhizosphere microbial communities is not known. To examine this question, we performed an experiment with barley (Hordeum vulgare) plants under iron-limiting and iron-sufficient growth conditions. Plants were grown in an iron-limiting soil in root box microcosms. One-half of the plants were treated with foliar iron every day to inhibit phytosiderophore production and to alter root exudate composition. After 30 days, the bacterial communities associated with different root zones, including the primary root tips, nonelongating secondary root tips, sites of lateral root emergence, and older roots distal from the tip, were characterized by using 16S ribosomal DNA (rDNA) fingerprints generated by PCR-denaturing gradient gel electrophoresis (DGGE). Our results showed that the microbial communities associated with the different root locations produced many common 16S rDNA bands but that the communities could be distinguished by using correspondence analysis. Approximately 40% of the variation between communities could be attributed to plant iron nutritional status. A sequence analysis of clones generated from a single 16S rDNA band obtained at all of the root locations revealed that there were taxonomically different species in the same band, suggesting that the resolving power of DGGE for characterization of community structure at the species level is limited. Our results suggest that the bacterial communities in the rhizosphere are substantially different in different root zones and that a rhizosphere community may be altered by changes in root exudate composition caused by changes in plant iron nutritional status.
Genetic Rearrangements of the Regions Adjacent to Genes Encoding Heat-Labile Enterotoxins (
eltAB
) of Enterotoxigenic
Escherichia coli
Strains
Authors: Schlör Stefan , Riedl Sabine , Blaß Julia , Reidl Joachim 
Published: April 2009

ABSTRACT

One of the most common bacterially mediated diarrheal infections is caused by enterotoxigenic
Escherichia coli
(ETEC) strains. ETEC-derived plasmids are responsible for the distribution of the genes encoding the main toxins, namely, the heat-labile and heat-stable enterotoxins. The origins and transfer modes (intra- or interplasmid) of the toxin-encoding genes have not been characterized in detail. In this study, we investigated the DNA regions located near the heat-labile enterotoxin-encoding genes (
eltAB
) of several clinical isolates. It was found that the
eltAB
region is flanked by conserved 236- and 280-bp regions, followed by highly variable DNA sequences which consist mainly of partial insertion sequence (IS) elements. Furthermore, we demonstrated that rearrangements of the
eltAB
region of one particular isolate, which harbors an IS
91
R sequence next to
eltAB
, could be produced by a
recA
-independent but IS
91
sequence-dependent mechanism. Possible mechanisms of dissemination of IS element-associated enterotoxin-encoding genes are discussed.
Use of Green Fluorescent Protein To Tag Lactic Acid Bacterium Strains under Development as Live Vaccine Vectors
Authors: Geoffroy Marie-Claude , Guyard Cyril , Quatannens Brigitte , Pavan Sonia , Lange Marc , Mercenier Annick 
Published: April 2009

ABSTRACT

The lactic acid bacteria (LAB) are safe microorganisms which are mainly used for the preparation of fermented foods and for probiotic applications. The potential of LAB as live vehicles for the production and delivery of therapeutic molecules such as antigens is also being actively investigated today. However, very little is known about the fate of live LAB when administered in vivo and about the interaction of these microorganisms with the nasal or gastrointestinal ecosystem. For future applications, it is essential to be able to discriminate the biotherapeutic strain from the endogenous microflora and to unravel the mechanisms underlying the postulated health-beneficial effect. We therefore started to investigate both aspects in a mouse model with two LAB species presently under development as live vaccine vectors, i.e.,
Lactococcus lactis
and
Lactobacillus plantarum
. We have constructed different expression vectors carrying the
gfp
(green fluorescent protein [GFP]) gene from the jellyfish
Aequoria victoria
, and we found that this visible marker was best expressed when placed under the control of the inducible strong
nisA
promoter from
L. lactis
. Notably, a threshold amount of GFP was necessary to obtain a bright fluorescent phenotype. We further demonstrated that fluorescent
L. plantarum
NCIMB8826 can be enumerated and sorted by flow cytometry. Moreover, tagging of this strain with GFP allowed us to visualize its phagocytosis by macrophages in vitro and ex vivo and to trace it in the gastrointestinal tract of mice upon oral administration.
Transcriptional Organization and Regulation of a Polycistronic Cold Shock Operon in
Sinorhizobium meliloti
RM1021 Encoding Homologs of the
Escherichia coli
Major Cold Shock Gene
cspA
and Ribosomal Protein Gene
rpsU
Authors: O'Connell Kevin P. , Thomashow Michael F. 
Published: April 2009

ABSTRACT

A homolog of the major eubacterial cold shock gene
cspA
was identified in
Sinorhizobium meliloti
RM1021 by
luxAB
reporter transposon mutagenesis. Here we further characterize the organization and regulation of this locus. DNA sequence analysis indicated that the locus includes three open reading frames (ORFs) encoding homologs corresponding to CspA, a novel 10.6-kDa polypeptide designated ORF2, and a homolog of the
Escherichia coli
ribosomal protein S21. Transcription analysis indicated that this locus produced two different-sized
cspA
-hybridizing transcripts upon cold shock, a 400-nucleotide (nt) RNA encoding
cspA
alone and a 1,000-nt transcript encoding
cspA-ORF2-rpsU
. The sizes of the transcripts agreed with the location of the transcription start site determined by primer extension and the locations of two putative transcriptional terminators. The promoter of the
cspA-ORF2-rpsU
locus had −10 and −35 elements similar to the
E. coli
ς
70
consensus promoter and, like the
cspA
locus of
E. coli
, included an AT-rich region upstream of the −35 hexamer. The promoter of the
S. meliloti cspA
locus was found to impart cold shock-induced mRNA accumulation. In addition, the 5′-untranslated region (5′ UTR) was found to increase the fold induction of
cspA
transcripts after cold shock and depressed the level of
luxAB
mRNA prior to cold shock, another feature similar to
cspA
regulation in
E. coli
. No “cold box” was identified upstream of the
S. meliloti cspA
gene, however, and there was no other obvious sequence identity between the
S. meliloti
5′ UTR and that of
E. coli
. DNA hybridization analysis indicated that outside the
cspA-ORF2-rpsU
cold shock locus there are several additional
cspA
-like genes and a second
rpsU
homolog.
Identification of Cold Shock Gene Loci in
Sinorhizobium meliloti
by Using a
luxAB
Reporter Transposon
Authors: O'Connell Kevin P. , Gustafson Ann M. , Lehmann M. Deane , Thomashow Michael F. 
Published: April 2009

ABSTRACT

Using a
luxAB
reporter transposon, seven mutants of
Sinorhizobium meliloti
were identified as containing insertions in four cold shock loci. LuxAB activity was strongly induced (25- to 160-fold) after a temperature shift from 30 to 15°C. The transposon and flanking host DNA from each mutant was cloned, and the nucleic acid sequence of the insertion site was determined. Unexpectedly, five of the seven
luxAB
mutants contained transposon insertions in the 16S and 23S rRNA genes of two of the three
rrn
operons of
S. meliloti
. Directed insertion of
luxAB
genes into each of the three
rrn
operons revealed that all three operons were similarly affected by cold shock. Two other insertions were found to be located downstream of a homolog of the major
Escherichia coli
cold shock gene,
cspA
. Although the cold shock loci were highly induced in response to a shift to low temperature, none of the insertions resulted in a statistically significant decrease in growth rate at 15°C.
Comparison of Animal Infectivity and Nucleic Acid Staining for Assessment of
Cryptosporidium parvum
Viability in Water
Authors: Neumann Norman F. , Gyürek Lyndon L. , Gammie Leslie , Finch Gordon R. , Belosevic Miodrag 
Published: April 2009

ABSTRACT

Cryptosporidium parvum
oocysts were stained with the fluorogenic dyes SYTO-9 and SYTO-59 and sorted by flow cytometry in order to determine whether the fluorescent staining intensity correlated with the ability of oocysts to infect neonatal CD-1 mice. Oocysts that did not fluoresce or that displayed weak fluorescent intensity when stained with SYTO-9 or SYTO-59 readily established infections in mice, whereas those oocysts that fluoresced brightly did not. Although fluorescent staining profiles varied among different batches of oocysts, a relative cutoff in fluorescent staining intensity that correlated with animal infectivity was observed for all batches.
Two Murine Monoclonal Antibodies against Serogroup E Salmonellae
Authors: Ng S. P. , Tsang R. S. W. , Luk J. M. C. , Ng M. H. , Im S. W. K. 
Published: April 2009

ABSTRACT

A monoclonal antibody (MAb), MO15, was raised against the lipopolysaccharide antigen of an ɛ15-lysogenized serogroup E
1
Salmonella
strain. The O factor 15-specific MAb MO15, together with another serogroup E-specific MAb, can differentiate among phage lysogenization variants in serogroup E salmonellae. Their epitope specificities in relation to conventional O-antigenic structures are discussed.