bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tissue Localization of the Endosymbiotic Bacterium “
Candidatus
Blochmannia floridanus” in Adults and Larvae of the Carpenter Ant
Camponotus floridanus
Authors: Sauer Christina , Dudaczek Dieter , Hölldobler Bert , Gross Roy 
Published: August 2002

ABSTRACT

The distribution of endosymbiotic bacteria in different tissues of queens, males, and workers of the carpenter ant
Camponotus floridanus
was investigated by light and electron microscopy and by in situ hybridization. A large number of bacteria could be detected in bacteriocytes within the midguts of workers, young virgin queens, and males. Large amounts of bacteria were also found in the oocytes of workers and queens. In contrast, bacteria were not present in oocyte-associated cells or in the spermathecae of mature queens, although occasionally a small number of bacteria could be detected in the testis follicles of males. Interestingly, the number of bacteriocytes in mature queens was strongly reduced and the bacteriocytes contained only very few or no bacteria at all, although the endosymbionts were present in huge amounts in the ovaries of the same animals. During embryogenesis of the deposited egg, the bacteria were concentrated in a ring of endodermal tissue destined to become the midgut in later developmental stages. However, during larval development, bacteria could also be detected in other tissues although to a lesser extent. Only in the last-instar larvae were bacteria found exclusively in the midgut tissue within typical bacteriocytes. Tetracycline and rifampin efficiently cleansed
C. floridanus
workers of their symbionts and the bacteriocytes of these animals still remained empty several months after treatment had ceased. Despite the lack of their endosymbionts, these adult animals were able to survive without any obvious negative effect under normal cultivation conditions.
Development of a Standard Test To Assess the Resistance of
Staphylococcus aureus
Biofilm Cells to Disinfectants
Authors: Luppens Suzanne B. I. , Reij Martine W. , van der Heijden Rob W. L. , Rombouts Frank M. , Abee Tjakko 
Published: August 2002

ABSTRACT

A standardized disinfectant test for
Staphylococcus aureus
cells in biofilms was developed. Two disinfectants, the membrane-active compound benzalkonium chloride (BAC) and the oxidizing agent sodium hypochlorite, were used to evaluate the biofilm test.
S. aureus
formed biofilms on glass, stainless steel, and polystyrene in a simple system with constant nutrient flow that mimicked as closely as possible the conditions used in the current standard European disinfectant test (EN 1040). The biofilm that was formed on glass contained cell clumps and extracellular polysaccharides. The average surface coverage was 60%, and most (92%) of the biofilm cells were viable. Biofilm formation and biofilm disinfection in different experiments were reproducible. For biofilms exposed to BAC and hypochlorite the concentrations needed to achieve 4-log killing were 50 and 600 times higher, respectively, than the concentrations needed to achieve this level of killing with the European phase 1 suspension test cells. Our results show that a standardized disinfectant test for biofilm cells is a useful addition to the current standard tests.
Effect of a
Sinorhizobium meliloti
Strain with a Modified
putA
Gene on the Rhizosphere Microbial Community of Alfalfa
Authors: van Dillewijn Pieter , Villadas Pablo J. , Toro Nicolás 
Published: August 2002

ABSTRACT

The success of a rhizobial inoculant in the soil depends to a large extent on its capacity to compete against indigenous strains. M403, a
Sinorhizobium meliloti
strain with enhanced competitiveness for nodule occupancy, was recently constructed by introducing a plasmid containing an extra copy of a modified
putA
(proline dehydrogenase) gene. This strain and M401, a control strain carrying the same plasmid without the modified gene, were used as soil inoculants for alfalfa in a contained field release experiment at León, Spain. In this study, we determined the effects of these two strains on the indigenous microbial community. 16S rRNA genes were obtained from the rhizosphere of alfalfa inoculated with strain M403 or strain M401 or from noninoculated plants by amplification of DNA from soil with bacterial group-specific primers. These genes were analyzed and compared by restriction fragment length polymorphism and temperature gradient gel electrophoresis. The results allowed us to differentiate between alterations in the microbial community apparently caused by inoculation and by the rhizosphere effect and seasonal fluctuations induced by the alfalfa plants and by the environment. Only moderate inoculation-dependent effects could be detected, while the alfalfa plants appeared to have a much stronger influence on the microbial community.
Environmental Regulation of
Campylobacter jejuni
Major Outer Membrane Protein Porin Expression in
Escherichia coli
Monitored by Using Green Fluorescent Protein
Authors: Dedieu Luc , Pagès Jean-Marie , Bolla Jean-Michel 
Published: August 2002

ABSTRACT

Porins allow exchanges between bacteria and their environment. In the gram-negative food-borne pathogen
Campylobacter jejuni
two porins, major outer membrane protein (MOMP) and Omp50, have been identified. MOMP is synthesized at a very high level under laboratory culture conditions, suggesting that its promoter functions very efficiently under these conditions. In
Campylobacter
samples, we observed that MOMP porin expression increased at a high temperature (42°C) or a high pH (pH 8.5) compared to expression at a low temperature (31°C) or an acidic pH (pH 5.5). To study the regulation of MOMP expression at the transcriptional level, we constructed an
momp-gfp
fusion in which
gfp
expression was put under the control of the
momp
promoter. Interestingly, we observed the same pattern of regulation in
Escherichia coli
, as monitored by green fluorescent protein production, that was found in
Campylobacter
. The ranges of pH and temperature tested are physiologically relevant, because they can be found in the digestive tracts of both birds and humans, which are both colonized by
Campylobacter.
Our results suggest that a component of the regulatory mechanism is conserved in
C. jejuni
and
E. coli
. However, medium osmolarity and sodium salicylate did not have a significant effect on
C. jejuni momp
promoter activity in
E. coli
, suggesting that major regulatory elements of
E. coli
porin expression do not participate in MOMP regulation. In contrast, mechanisms involving DNA supercoiling may be involved, as shown by DNA gyrase inhibition assays. These findings are a step towards determining the role of outer membrane proteins in the adaptation of
C. jejuni
to its environment.
Genetic Heterogeneity in
Bacillus sporothermodurans
as Demonstrated by Ribotyping and Repetitive Extragenic Palindromic-PCR Fingerprinting
Authors: Guillaume-Gentil Olivier , Scheldeman Patsy , Marugg Joey , Herman Lieve , Joosten Han , Heyndrickx Marc 
Published: August 2002

ABSTRACT

Thirty-eight strains of
Bacillus sporothermodurans
isolated from ultra-high-temperature (UHT)-treated milk or sterilized milk (UHT isolates) and from animal feed or raw milk (farm isolates) were characterized by automated ribotyping and by repetitive extragenic palindromic (REP)-PCR fingerprinting. By investigating the genetic relationships among isolates from these various sources, the relative importance of different contamination sources could be evaluated. The results of the separate clustering analyses of the
Pvu
II and
Eco
RI ribopatterns and the REP-PCR patterns were largely consistent with each other and revealed the existence of two main clusters; there was one homogeneous group containing all (REP-PCR) or most (ribotyping) of the UHT isolates, and there was a second more diverse group comprising the farm isolates. A combined three-dimensional analysis of all data showed that three German UHT isolates did not belong to the compact group containing the majority of the UHT isolates. These results demonstrate that
B. sporothermodurans
is more heterogeneous than previously assumed and that most of the UHT isolates form a genetically distinct subgroup and are capable of producing highly heat-resistant spores. The close genetic relationship of these UHT isolates suggests a clonal origin of a few predominant strains of
B. sporothermodurans
that can be found in UHT-treated or sterilized milk products.
Quantification of Uncultured
Ruminococcus obeum
-Like Bacteria in Human Fecal Samples by Fluorescent In Situ Hybridization and Flow Cytometry Using 16S rRNA-Targeted Probes
Authors: Zoetendal Erwin G. , Ben-Amor Kaouther , Harmsen Hermie J. M. , Schut Frits , Akkermans Antoon D. L. , de Vos Willem M. 
Published: August 2002

ABSTRACT

A 16S rRNA-targeted probe was designed and validated in order to quantify the number of uncultured
Ruminococcus obeum
-like bacteria by fluorescent in situ hybridization (FISH). These bacteria have frequently been found in 16S ribosomal DNA clone libraries prepared from bacterial communities in the human intestine. Thirty-two reference strains from the human intestine, including a phylogenetically related strain and strains of some other
Ruminococcus
species, were used as negative controls and did not hybridize with the new probe. Microscopic and flow cytometric analyses revealed that a group of morphologically similar bacteria in feces did hybridize with this probe. Moreover, it was found that all hybridizing cells also hybridized with a probe specific for the
Clostridium coccoides-Eubacterium rectale
group, a group that includes the uncultured
R. obeum
-like bacteria. Quantification of the uncultured
R. obeum
-like bacteria and the
C. coccoides-E. rectale
group by flow cytometry and microscopy revealed that these groups comprised approximately 2.5 and 16% of the total community in fecal samples, respectively. The uncultured
R. obeum
-like bacteria comprise about 16% of the
C. coccoides-E. rectale
group. These results indicate that the uncultured
R. obeum
-like bacteria are numerically important in human feces. Statistical analysis revealed no significant difference between the microscopic and flow cytometric counts and the different feces sampling times, while a significant host-specific effect on the counts was observed. Our data demonstrate that the combination of FISH and flow cytometry is a useful approach for studying the ecology of uncultured bacteria in the human gastrointestinal tract.
Cyclization Reaction Catalyzed by Glycogen Debranching Enzyme (EC 2.4.1.25/EC 3.2.1.33) and Its Potential for Cycloamylose Production
Authors: Yanase Michiyo , Takata Hiroki , Takaha Takeshi , Kuriki Takashi , Smith Steven M. , Okada Shigetaka 
Published: August 2002

ABSTRACT

Glycogen debranching enzyme (GDE) has 4-α-glucanotransferase and amylo-1,6-glucosidase activities in the single polypeptide chain. We analyzed the detailed action profile of GDE from
Saccharomyces cerevisiae
on amylose and tested whether GDE catalyzes cyclization of amylose. GDE treatment resulted in a rapid reduction of absorbance of iodine-amylose complex and the accumulation of a product that was resistant to an exo-amylase (glucoamylase [GA]) but was degraded by an endo-type α-amylase to glucose and maltose. These results indicated that GDE catalyzed cyclization of amylose to produce cyclic α-1,4 glucan (cycloamylose). The formation of cycloamylose was confirmed by high-performance anion-exchange chromatography, and the size was shown to range from a degree of polymerization of 11 to a degree of polymerization around 50. The minimum size and the size distribution of cycloamylose were different from those of cycloamylose produced by other 4-α-glucanotransferases. GDE also efficiently produced cycloamylose even from the branched glucan substrate, starch, demonstrating its potential for industrial production of cycloamylose.
A Stationary-Phase Acyl-Coenzyme A Synthetase of
Streptomyces coelicolor
A3(2) Is Necessary for the Normal Onset of Antibiotic Production
Authors: Banchio C. , Gramajo H. 
Published: August 2002

ABSTRACT

The
fadD1
and
macs1
genes of
Streptomyces coelicolor
are part of a two-gene operon. Both genes encode putative acyl coenzyme A synthetases (ACSs). The amino acid sequence of FadD1 has high homology with those of several ACSs, while MACS1 has the closest homology with medium-chain ACSs, broadly known as SA proteins. Like FadD of
Escherichia coli
, FadD1 also has a broad substrate specificity, although saturated long-chain fatty acids appears to be the preferred substrate.
fadD1
is a growth-phase-regulated gene, and its mRNA is detected only during the stationary phase of growth. Interestingly, a mutation in
fadD1
alters the levels of another ACS or ACSs, both at the stationary phase and at the exponential phase of growth, at least when glucose is used as a main carbon source. The mutant also shows a severe deficiency in antibiotic production, and at least for Act biosynthesis, this deficiency seems to be related to delayed expression of the Act biosynthetic genes. Antibiotic production is restored by the introduction of a wt
fadD1
allele into the cell, demonstrating a strict link between ACS activity and the biosynthesis of secondary metabolites. The results of this study indicate that the ACSs may be useful targets for the design of rational approaches to improving antibiotic production.
Biochemical Evidence for Formate Transfer in Syntrophic Propionate-Oxidizing Cocultures of
Syntrophobacter fumaroxidans
and
Methanospirillum hungatei
Authors: de Bok Frank A. M. , Luijten Maurice L. G. C. , Stams Alfons J. M. 
Published: August 2002

ABSTRACT

The hydrogenase and formate dehydrogenase levels in
Syntrophobacter fumaroxidans
and
Methanospirillum hungatei
were studied in syntrophic propionate-oxidizing cultures and compared to the levels in axenic cultures of both organisms. Cells grown syntrophically were separated from each other by Percoll gradient centrifugation. In
S. fumaroxidans
both formate dehydrogenase and hydrogenase levels were highest in cells which were grown syntrophically, while the formate-H
2
lyase activities were comparable under the conditions tested. In
M. hungatei
the formate dehydrogenase and formate-H
2
lyase levels were highest in cells grown syntrophically, while the hydrogenase levels in syntrophically grown cells were comparable to those in cells grown on formate. Reconstituted syntrophic cultures from axenic cultures immediately resumed syntrophic growth, and the calculated growth rates of these cultures were highest for cells which were inoculated from the axenic
S. fumaroxidans
cultures that exhibited the highest formate dehydrogenase activities. The results suggest that formate is the preferred electron carrier in syntrophic propionate-oxidizing cocultures of
S. fumaroxidans
and
M. hungatei
.
Purification and Immunogenicity of a RecombinantBordetella pertussisS1S3FHA Fusion Protein Expressed byStreptococcus gordonii
Authors: Lee Song F. , Halperin Scott A. , Knight Jennifer B. , Tait Aaron 
Published: August 2002

ABSTRACTAcellular pertussis vaccines typically consist of antigens isolated fromBordetella pertussis, and pertussis toxin (PT) and filamentous hemagglutinin (FHA) are two prominent components. One of the disadvantages of a multiple-component vaccine is the cost associated with the production of the individual components. In this study, we constructed an in-frame fusion protein consisting of PT fragments (179 amino acids of PT subunit S1 and 180 amino acids of PT subunit S3) and a 456-amino-acid type I domain of FHA. The fusion protein was expressed by the commensal oral bacteriumStreptococcus gordonii.The fusion protein was secreted into the culture medium as an expected 155-kDa protein, which was recognized by a polyclonal anti-PT antibody, a monoclonal anti-S1 antibody, and a monoclonal anti-FHA antibody. The fusion protein was purified from the culture supernatant by affinity and gel permeation chromatography. The immunogenicity of the purified fusion protein was assessed in BALB/c mice by performing parenteral and mucosal immunization experiments. When given parenterally, the fusion protein elicited a very strong antibody titer against the FHA type I domain, a moderate titer against native FHA, and a weak titer against PT. When given mucosally, it elicited a systemic response and a mucosal response to FHA and PT. In Western blots, the immune sera recognized the S1, S3, and S2 subunits of PT. These data collectively indicate that fragments of the pertussis vaccine components can be expressed in a single fusion protein byS. gordoniiand that the fusion protein is immunogenic. This multivalent fusion protein approach may be used in designing a new generation of acellular pertussis vaccines.
Hxt-Carrier-Mediated Glucose Efflux upon Exposure of
Saccharomyces cerevisiae
to Excess Maltose
Authors: Jansen Mickel L. A. , De Winde Johannes H. , Pronk Jack T. 
Published: August 2002

ABSTRACT

When wild-type
Saccharomyces cerevisiae
strains pregrown in maltose-limited chemostat cultures were exposed to excess maltose, release of glucose into the external medium was observed. Control experiments confirmed that glucose release was not caused by cell lysis or extracellular maltose hydrolysis. To test the hypothesis that glucose efflux involved plasma membrane glucose transporters, experiments were performed with an
S. cerevisiae
strain in which all members of the hexose transporter (
HXT
) gene family had been eliminated and with an isogenic reference strain. Glucose efflux was virtually eliminated in the hexose-transport-deficient strain. This constitutes experimental proof that Hxt transporters facilitate export of glucose from
S. cerevisiae
cells. After exposure of the hexose-transport-deficient strain to excess maltose, an increase in the intracellular glucose level was observed, while the concentrations of glucose 6-phosphate and ATP remained relatively low. These results demonstrate that glucose efflux can occur as a result of uncoordinated expression of the initial steps of maltose metabolism and the subsequent reactions in glucose dissimilation. This is a relevant phenomenon for selection of maltose-constitutive strains for baking and brewing.
Albidovulum inexpectatum
gen. nov., sp. nov., a Nonphotosynthetic and Slightly Thermophilic Bacterium from a Marine Hot Spring That Is Very Closely Related to Members of the Photosynthetic Genus
Rhodovulum
Authors: Albuquerque Luciana , Santos João , Travassos Pedro , Nobre M. Fernanda , Rainey Fred A. , Wait Robin , Empadinhas Nuno , Silva Manuel T. , da Costa Milton S. 
Published: August 2002

ABSTRACT

Several bacterial isolates, with an optimum growth temperature of about 50°C, were recovered from the marine hot spring at Ferraria on the island of São Miguel in the Azores. The geothermal water emerged from a porous lava flow and rapidly cooled in contact with seawater except at low tide. The bacterial species represented by strains FRR-10
T
and FRR-11 was nonpigmented, strictly aerobic, and organotrophic. Several genes,
bchZ
,
pufB
,
pufA
,
pufL
, or
pufM
, encoding the photosynthetic reaction center proteins and the core light-harvesting complexes were not detected in these strains. The organism oxidized thiosulfate to sulfate with enhancement of growth. The organism did not require additional NaCl in the culture medium for growth, but NaCl at 1.0% enhanced growth. Phylogenetic analyses using the 16S rRNA gene sequence of strain FRR-10
T
indicated that the new organism represented a new species of the α-3 subclass of the
Proteobacteria
and that it branches within the species of the genus
Rhodovulum
. The contradiction of classifying an organism which branches within the radiation of the genus
Rhodovulum
but does not possess the hallmark characteristics of this genus is discussed. However, the absence of several of these characteristics, namely, the lack of photosynthesis and pigmentation, which could be related to colonization of dark environments, and growth at high temperatures, leads to our proposal that strains FRR-10
T
and FRR-11 should be classified as a new species of a novel genus,
Albidovulum inexpectatum
, representing, at present, the most thermophilic organism within the α-3 subclass of the
Proteobacteria
.
Expression of Genes Encoding F
1
-ATPase Results in Uncoupling of Glycolysis from Biomass Production in
Lactococcus lactis
Authors: Koebmann Brian J. , Solem Christian , Pedersen Martin B. , Nilsson Dan , Jensen Peter R. 
Published: August 2002

ABSTRACT

We studied how the introduction of an additional ATP-consuming reaction affects the metabolic fluxes in
Lactococcus lactis
. Genes encoding the hydrolytic part of the F
1
domain of the membrane-bound (F
1
F
0
) H
+
-ATPase were expressed from a range of synthetic constitutive promoters. Expression of the genes encoding F
1
-ATPase was found to decrease the intracellular energy level and resulted in a decrease in the growth rate. The yield of biomass also decreased, which showed that the incorporated F
1
-ATPase activity caused glycolysis to be uncoupled from biomass production. The increase in ATPase activity did not shift metabolism from homolactic to mixed-acid fermentation, which indicated that a low energy state is not the signal for such a change. The effect of uncoupled ATPase activity on the glycolytic flux depended on the growth conditions. The uncoupling stimulated the glycolytic flux threefold in nongrowing cells resuspended in buffer, but in steadily growing cells no increase in flux was observed. The latter result shows that glycolysis occurs close to its maximal capacity and indicates that control of the glycolytic flux under these conditions resides in the glycolytic reactions or in sugar transport.
Molecular Characterization of a Novel Glucosyltransferase from
Lactobacillus reuteri
Strain 121 Synthesizing a Unique, Highly Branched Glucan with α-(1→4) and α-(1→6) Glucosidic Bonds
Authors: Kralj S. , van Geel-Schutten G. H. , Rahaoui H. , Leer R. J. , Faber E. J. , van der Maarel M. J. E. C. , Dijkhuizen L. 
Published: August 2002

ABSTRACT

Lactobacillus reuteri
strain 121 produces a unique, highly branched, soluble glucan in which the majority of the linkages are of the α-(1→4) glucosidic type. The glucan also contains α-(1→6)-linked glucosyl units and 4,6-disubstituted α-glucosyl units at the branching points. Using degenerate primers, based on the amino acid sequences of conserved regions from known glucosyltransferase (
gtf
) genes from lactic acid bacteria, the
L. reuteri
strain 121 glucosyltransferase gene (
gtfA
) was isolated. The
gtfA
open reading frame (ORF) was 5,343 bp, and it encodes a protein of 1,781 amino acids with a deduced
M
r
of 198,637. The deduced amino acid sequence of GTFA revealed clear similarities with other glucosyltransferases. GTFA has a relatively large variable N-terminal domain (702 amino acids) with five unique repeats and a relatively short C-terminal domain (267 amino acids). The
gtfA
gene was expressed in
Escherichia coli
, yielding an active GTFA enzyme. With respect to binding type and size distribution, the recombinant GTFA enzyme and the
L. reuteri
strain 121 culture supernatants synthesized identical glucan polymers. Furthermore, the deduced amino acid sequence of the
gtfA
ORF and the N-terminal amino acid sequence of the glucosyltransferase isolated from culture supernatants of
L. reuteri
strain 121 were the same. GTFA is thus responsible for the synthesis of the unique glucan polymer in
L. reuteri
strain 121. This is the first report on the molecular characterization of a glucosyltransferase from a
Lactobacillus
strain.