bluuebunny/crossref_metadata_2025_split huggingface dataset:
How Clonal Is
Staphylococcus aureus
?
Authors: Feil Edward J. , Cooper Jessica E. , Grundmann Hajo , Robinson D. Ashley , Enright Mark C. , Berendt Tony , Peacock Sharon J. , Smith John Maynard , Murphy Michael , Spratt Brian G. , Moore Catrin E. , Day Nicholas P. J. 
Published: May 2003

ABSTRACT

Staphylococcus aureus
is an important human pathogen and represents a growing public health burden owing to the emergence and spread of antibiotic-resistant clones, particularly within the hospital environment. Despite this, basic questions about the evolution and population biology of the species, particularly with regard to the extent and impact of homologous recombination, remain unanswered. We address these issues through an analysis of sequence data obtained from the characterization by multilocus sequence typing (MLST) of 334 isolates of
S
.
aureus
, recovered from a well-defined population, over a limited time span. We find no significant differences in the distribution of multilocus genotypes between strains isolated from carriers and those from patients with invasive disease; there is, therefore, no evidence from MLST data, which index variation within the stable “core” genome, for the existence of hypervirulent clones of this pathogen. Examination of the sequence changes at MLST loci during clonal diversification shows that point mutations give rise to new alleles at least 15-fold more frequently than does recombination. This contrasts with the naturally transformable species
Neisseria meningitidis
and
Streptococcus pneumoniae
, in which alleles change between 5- and 10-fold more frequently by recombination than by mutation. However, phylogenetic analysis suggests that homologous recombination does contribute toward the evolution of this species over the long term. Finally, we note a striking excess of nonsynonymous substitutions in comparisons between isolates belonging to the same clonal complex compared to isolates belonging to different clonal complexes, suggesting that the removal of deleterious mutations by purifying selection may be relatively slow.
Novel Developmental Genes,
fruCD
, of
Myxococcus xanthus
: Involvement of a Cell Division Protein in Multicellular Development
Authors: Akiyama Takuya , Inouye Sumiko , Komano Teruya 
Published: May 2003

ABSTRACT

Myxococcus xanthus
is a gram-negative soil bacterium that undergoes multicellular development upon nutrient starvation. In the present study, two novel developmental genes,
fruC
and
fruD
, of
M. xanthus
were identified and characterized. The FruD protein has significant amino acid sequence similarity to the DivIVA proteins of many bacteria including
Bacillus subtilis
. Vegetative cells of the
fruD
mutant exhibited a filamentous phenotype. The
fruC
and
fruD
mutants displayed similar delayed-development phenotypes. The formation of tightly aggregated mounds by
fruC
and
fruD
mutants was slower than that by the wild-type strain. Spore formation by the
fruC
and
fruD
mutants initiated after 30 h poststarvation, whereas wild-type
M. xanthus
initiated spore formation after 18 h. The
fruCD
genes were constitutively expressed as an operon during vegetative growth and development. S1 mapping revealed that transcription initiation sites of the
fruCD
operon were located 114 (P1) and 55 bp (P2) upstream of the
fruC
initiation codon. Only the P1 promoter was active during vegetative growth, while both the P1 and P2 promoters were active during development. The FruD protein was produced as a cytoplasmic protein and formed an oligomer during vegetative growth and development.
Genomic Sequence of C
1
, the First Streptococcal Phage
Authors: Nelson Daniel , Schuch Raymond , Zhu Shiwei , Tscherne Donna M. , Fischetti Vincent A. 
Published: May 2003

ABSTRACT

C
1
, a lytic bacteriophage infecting group C streptococci, is one of the earliest-isolated phages, and the method of bacterial classification known as phage typing was defined by using this bacteriophage. We present for the first time a detailed analysis of this phage by use of electron microscopy, protein profiling, and complete nucleotide sequencing. This virus belongs to the
Podoviridae
family of phages, all of which are characterized by short, noncontractile tails. The C
1
genome consists of a linear double-stranded DNA molecule of 16,687 nucleotides with 143-bp inverted terminal repeats. We have assigned functions to 9 of 20 putative open reading frames based on experimental substantiation or bioinformatic analysis. Their products include DNA polymerase, holin, lysin, major capsid, head-tail connector, neck appendage, and major tail proteins. Additionally, we found one intron belonging to the HNH endonuclease family interrupting the apparent lysin gene, suggesting a potential splicing event yielding a functional lytic enzyme. Examination of the C
1
DNA polymerase suggests that this phage utilizes a protein-primed mechanism of replication, which is prominent in the φ29-like members of
Podoviridae
. Consistent with this evidence, we experimentally determined that terminal proteins are covalently attached to both 5′ termini, despite the fact that no homology to known terminal proteins could be elucidated in any of our open reading frames. Likewise, comparative genomics revealed no close evolutionary matches, suggesting that the C
1
bacteriophage is a unique member of the
Podoviridae
.
Distribution and Organization of Auxotrophic Genes on the Multichromosomal Genome of
Burkholderia multivorans
ATCC 17616
Authors: Komatsu Harunobu , Imura Yoshiyuki , Ohori Akira , Nagata Yuji , Tsuda Masataka 
Published: May 2003

ABSTRACT

The
Burkholderia multivorans
strain ATCC 17616 carries three circular chromosomes with sizes of 3.4, 2.5, and 0.9 Mb. To determine the distribution and organization of the amino acid biosynthetic genes on the genome of this β-proteobacterium, various auxotrophic mutations were isolated using a Tn
5
derivative that was convenient not only for the determination of its insertion site on the genome map but also for the structural analysis of the flanking regions. Analysis by pulsed-field gel electrophoresis revealed that 20 out of 23 insertion mutations were distributed on the 3.4-Mb chromosome. More detailed analysis of the
his
,
trp
,
arg
, and
lys
mutations and their flanking regions revealed the following properties of these auxotrophic genes: (i) all nine
his
genes were clustered on the 3.4-Mb chromosome; (ii) seven
trp
genes were organized within two distinct regions, i.e., a
trpEGDC
cluster on the 3.4-Mb chromosome and a
trpFBA
cluster on the 2.5-Mb chromosome; (iii) the
leu
gene cluster,
leuCDB
, was also located close to the
trpFBA
cluster; and (iv)
lysA
and
argG
genes were located on the 2.5-Mb chromosome, in contrast to the
argH
gene, which was located on the 3.4-Mb chromosome. Southern hybridization analysis, allelic exchange mutagenesis of ATCC 17616, and complementation tests demonstrated that all of the genes examined were functional and existed as a single copy within the genome. The present findings also indicated that the 2.5-Mb chromosome carried various auxotrophic genes with no structural or functional counterparts on the remaining two chromosomes.
Concentration and Assembly of the Division Ring Proteins FtsZ, FtsA, and ZipA during theEscherichia coliCell Cycle
Authors: Rueda Sonsoles , Vicente Miguel , Mingorance Jesús 
Published: May 2003

ABSTRACTThe concentration of the cell division proteins FtsZ, FtsA, and ZipA and their assembly into a division ring during theEscherichia coliB/r K cell cycle have been measured in synchronous cultures obtained by the membrane elution technique. Immunostaining of the three proteins revealed no organized structure in newly born cells. In a culture with a doubling time of 49 min, assembly of the Z ring started around minute 25 and was detected first as a two-dot structure that became a sharp band before cell constriction. FtsA and ZipA localized into a division ring following the same pattern and time course as FtsZ. The concentration (amount relative to total mass) of the three proteins remained constant during one complete cell cycle, showing that assembly of a division ring is not driven by changes in the concentration of these proteins. Maintenance of the Z ring during the process of septation is a dynamic energy-dependent event, as evidenced by its disappearance in cells treated with sodium azide.
Genomic Analysis and Initial Characterization of the Chitinolytic System of
Microbulbifer degradans
Strain 2-40
Authors: Howard Michael B. , Ekborg Nathan A. , Taylor Larry E. , Weiner Ronald M. , Hutcheson Steven W. 
Published: May 2003

ABSTRACT

The marine bacterium
Microbulbifer degradans
strain 2-40 produces at least 10 enzyme systems for degrading insoluble complex polysaccharides (ICP). The draft sequence of the 2-40 genome allowed a genome-wide analysis of the chitinolytic system of strain 2-40. The chitinolytic system includes three secreted chitin depolymerases (ChiA, ChiB, and ChiC), a secreted chitin-binding protein (CbpA), periplasmic chitooligosaccharide-modifying enzymes, putative sugar transporters, and a cluster of genes encoding cytoplasmic proteins involved in
N
-acetyl-
d
-glucosamine (GlcNAc) metabolism. Each chitin depolymerase was detected in culture supernatants of chitin-grown strain 2-40 and was active against chitin and glycol chitin. The chitin depolymerases also had a specific pattern of activity toward the chitin analogs 4-methylumbelliferyl-β-
d
-
N
,
N
′-diacetylchitobioside (MUF-diNAG) and 4-methylumbelliferyl-β-
d
-
N
,
N
′,
N
"-triacetylchitotrioside (MUF-triNAG). The depolymerases were modular in nature and contained glycosyl hydrolase family 18 domains, chitin-binding domains, and polycystic kidney disease domains. ChiA and ChiB each possessed polyserine linkers of up to 32 consecutive serine residues. In addition, ChiB and CbpA contained glutamic acid-rich domains. At 1,271 amino acids, ChiB is the largest bacterial chitinase reported to date. A chitodextrinase (CdxA) with activity against chitooligosaccharides (degree of polymerization of 5 to 7) was identified. The activities of two apparent periplasmic (HexA and HexB)
N
-acetyl-β-
d
-glucosaminidases and one cytoplasmic (HexC)
N
-acetyl-β-
d
-glucosaminidase were demonstrated. Genes involved in GlcNAc metabolism, similar to those of the
Escherichia coli
K-12 NAG utilization operon, were identified. NagA from strain 2-40, a GlcNAc deacetylase, was shown to complement a
nagA
mutation in
E
.
coli
K-12. Except for the GlcNAc utilization cluster, genes for all other components of the chitinolytic system were dispersed throughout the genome. Further examination of this system may provide additional insight into the mechanisms by which marine bacteria degrade chitin and provide a basis for future research on the ICP-degrading systems of strain 2-40.
The Dithiol:Disulfide Oxidoreductases DsbA and DsbB of
Rhodobacter capsulatus
Are Not Directly Involved in Cytochrome
c
Biogenesis, but Their Inactivation Restores the Cytochrome
c
Biogenesis Defect of CcdA-Null Mutants
Authors: Deshmukh Meenal , Turkarslan Serdar , Astor Donniel , Valkova-Valchanova Maria , Daldal Fevzi 
Published: May 2003

ABSTRACT

The cytoplasmic membrane protein CcdA and its homologues in other species, such as DsbD of
Escherichia coli
, are thought to supply the reducing equivalents required for the biogenesis of
c
-type cytochromes that occurs in the periplasm of gram-negative bacteria. CcdA
-
null mutants of the facultative phototroph
Rhodobacter capsulatus
are unable to grow under photosynthetic conditions (Ps
−
) and do not produce any active cytochrome
c
oxidase (Nadi
−
) due to a pleiotropic cytochrome
c
deficiency. However, under photosynthetic or respiratory growth conditions, these mutants revert frequently to yield Ps
+
Nadi
+
colonies that produce
c
-type cytochromes despite the absence of CcdA. Complementation of a CcdA-null mutant for the Ps
+
growth phenotype was attempted by using a genomic library constructed with chromosomal DNA from a revertant. No complementation was observed, but plasmids that rescued a CcdA-null mutant for photosynthetic growth by homologous recombination were recovered. Analysis of one such plasmid revealed that the rescue ability was mediated by open reading frame 3149, encoding the dithiol:disulfide oxidoreductase DsbA. DNA sequence data revealed that the
dsbA
allele on the rescuing plasmid contained a frameshift mutation expected to produce a truncated, nonfunctional DsbA. Indeed, a
dsbA ccdA
double mutant was shown to be Ps
+
Nadi
+
, establishing that in
R. capsulatus
the inactivation of
dsbA
suppresses the
c
-type cytochrome deficiency due to the absence of
ccdA
. Next, the ability of the wild-type
dsbA
allele to suppress the Ps
+
growth phenotype of the
dsbA ccdA
double mutant was exploited to isolate
dsbA
-independent
ccdA
revertants. Sequence analysis revealed that these revertants carried mutations in
dsbB
and that their Ps
+
phenotypes could be suppressed by the wild-type allele of
dsbB
. As with
dsbA
, a
dsbB ccdA
double mutant was also Ps
+
Nadi
+
and produced
c
-type cytochromes. Therefore, the absence of either DsbA or DsbB restores
c
-type cytochrome biogenesis in the absence of CcdA. Finally, it was also found that the DsbA-null and DsbB-null single mutants of
R. capsulatus
are Ps
+
and produce
c
-type cytochromes, unlike their
E. coli
counterparts, but are impaired for growth under respiratory conditions. This finding demonstrates that in
R. capsulatus
the dithiol:disulfide oxidoreductases DsbA and DsbB are not essential for cytochrome
c
biogenesis even though they are important for respiration under certain conditions.
Genes of
Bacillus cereus
and
Bacillus anthracis
Encoding Proteins of the Exosporium
Authors: Todd Sarah J. , Moir Arthur J. G. , Johnson Matt J. , Moir Anne 
Published: May 2003

ABSTRACT

The exosporium is the outermost layer of spores of
Bacillus cereus
and its close relatives
Bacillus anthracis
and
Bacillus thuringiensis
. For these pathogens, it represents the surface layer that makes initial contact with the host. To date, only the BclA glycoprotein has been described as a component of the exosporium; this paper defines 10 more tightly associated proteins from the exosporium of
B. cereus
ATCC 10876, identified by N-terminal sequencing of proteins from purified, washed exosporium. Likely coding sequences were identified from the incomplete genome sequence of
B. anthracis
or
B. cereus
ATCC 14579, and the precise corresponding sequence from
B. cereus
ATCC 10876 was defined by PCR and sequencing. Eight genes encode likely structural components (
exsB
,
exsC
,
exsD
,
exsE
,
exsF
,
exsG
,
exsJ
, and
cotE
). Several proteins of the exosporium are related to morphogenetic and outer spore coat proteins of
B. subtilis
, but most do not have homologues in
B. subtilis
. ExsE is processed from a larger precursor, and the CotE homologue appears to have been C-terminally truncated. ExsJ contains a domain of GXX collagen-like repeats, like the BclA exosporium protein of
B. anthracis
. Although most of the exosporium genes are scattered on the genome,
bclA
and
exsF
are clustered in a region flanking the rhamnose biosynthesis operon; rhamnose is part of the sugar moiety of spore glycoproteins. Two enzymes, alanine racemase and nucleoside hydrolase, are tightly adsorbed to the exosporium layer; they could metabolize small molecule germinants and may reduce the sensitivity of spores to these, limiting premature germination.
Sigma 54 Levels and Physiological Control of the
Pseudomonas putida
Pu Promoter
Authors: Jurado Paola , Fernández Luis A. , de Lorenzo Víctor 
Published: May 2003

ABSTRACT

The cellular levels of the alternative sigma factor σ
54
of
Pseudomonas putida
have been examined in a variety of growth stages and culture conditions with a single-chain Fv antibody tailored for detection of scarce proteins. The levels of σ
54
were also monitored in
P. putida
strains with knockout mutations in
ptsO
or
ptsN
, known to be required for the C-source control of the σ
54
-dependent Pu promoter of the TOL plasmid. Our results show that ∼80 ± 26 molecules of σ
54
exist per cell. Unlike that in relatives of
Pseudomonas
(e.g.,
Caulobacter
), where fluctuations of σ
54
determine adaptation and differentiation when cells face starvation, σ
54
in
P. putida
remains unexpectedly constant at different growth stages, in nitrogen starvation and C-source repression conditions, and in the
ptsO
and
ptsN
mutant strains analyzed. The number of σ
54
molecules per cell in
P. putida
is barely above the predicted number of σ
54
-dependent promoters. These figures impose a framework on the mechanism by which Pu (and other σ
54
-dependent systems) may become amenable to physiological control.
Polar Localization of Replicon Origins in the Multipartite Genomes ofAgrobacterium tumefaciensandSinorhizobium meliloti
Authors: Kahng Lyn Sue , Shapiro Lucy 
Published: May 2003

ABSTRACTThe origins of replication of many different bacteria have been shown to reside at specific subcellular locations, but the mechanisms underlying their positioning and segregation are still being elucidated. In particular, little is known about the replication of multipartite genomes in bacteria. We determined the cellular positions of the origins of the replicons in the alpha proteobacteriaAgrobacterium tumefaciensandSinorhizobium melilotiand found that they are located at the poles of the cells. Our work demonstrates the conserved extreme polar localization of circular chromosome origins in these alpha proteobacteria and is also the first to specify the cellular location of origin regions from therepABCfamily. The cellular location of a derivative of the RK2 plasmid is distinct from that of the alpha proteobacterium genomic replicon origins but is conserved across bacteria. Colocalization experiments with the genomic replicons ofA. tumefaciensrevealed that therepABCreplicons, although preferentially positioned at the cell pole, colocalize only rarely. For therepABCreplicons in this organism, occupying discrete spatial locations may contribute to their coexistence and stable inheritance.
Modeling Bacterial Evolution with Comparative-Genome-Based Marker Systems: Application to
Mycobacterium tuberculosis
Evolution and Pathogenesis
Authors: Alland David , Whittam Thomas S. , Murray Megan B. , Cave M. Donald , Hazbon Manzour H. , Dix Kim , Kokoris Mark , Duesterhoeft Andreas , Eisen Jonathan A. , Fraser Claire M. , Fleischmann Robert D. 
Published: May 2003

ABSTRACT

The comparative-genomic sequencing of two
Mycobacterium tuberculosis
strains enabled us to identify single nucleotide polymorphism (SNP) markers for studies of evolution, pathogenesis, and epidemiology in clinical
M
.
tuberculosis
. Phylogenetic analysis using these “comparative-genome markers” (CGMs) produced a highly unusual phylogeny with a complete absence of secondary branches. To investigate CGM-based phylogenies, we devised computer models to simulate sequence evolution and calculate new phylogenies based on an SNP format. We found that CGMs represent a distinct class of phylogenetic markers that depend critically on the genetic distances between compared “reference strains.” Properly distanced reference strains generate CGMs that accurately depict evolutionary relationships, distorted only by branch collapse. Improperly distanced reference strains generate CGMs that distort and reroot outgroups. Applying this understanding to the CGM-based phylogeny of
M
.
tuberculosis
, we found evidence to suggest that this species is highly clonal without detectable lateral gene exchange. We noted indications of evolutionary bottlenecks, including one at the level of the PHRI “C” strain previously associated with particular virulence characteristics. Our evidence also suggests that loss of IS
6110
to fewer than seven elements per genome is uncommon. Finally, we present population-based evidence that KasA, an important component of mycolic acid biosynthesis, develops G312S polymorphisms under selective pressure.
Coaggregation-Mediated Interactions of Streptococci and Actinomyces Detected in Initial Human Dental Plaque
Authors: Palmer, Robert J. , Gordon Sharon M. , Cisar John O. , Kolenbrander Paul E. 
Published: May 2003

ABSTRACT

Streptococci and actinomyces that initiate colonization of the tooth surface frequently coaggregate with each other as well as with other oral bacteria. These observations have led to the hypothesis that interbacterial adhesion influences spatiotemporal development of plaque. To assess the role of such interactions in oral biofilm formation in vivo, antibodies directed against bacterial surface components that mediate coaggregation interactions were used as direct immunofluorescent probes in conjunction with laser confocal microscopy to determine the distribution and spatial arrangement of bacteria within intact human plaque formed on retrievable enamel chips. In intrageneric coaggregation, streptococci such as
Streptococcus gordonii
DL1 recognize receptor polysaccharides (RPS) borne on other streptococci such as
Streptococcus oralis
34. To define potentially interactive subsets of streptococci in the developing plaque, an antibody against RPS (anti-RPS) was used together with an antibody against
S. gordonii
DL1 (anti-DL1). These antibodies reacted primarily with single cells in 4-h-old plaque and with mixed-species microcolonies in 8-h-old plaque. Anti-RPS-reactive bacteria frequently formed microcolonies with anti-DL1-reactive bacteria and with other bacteria distinguished by general nucleic acid stains. In intergeneric coaggregation between streptococci and actinomyces, type 2 fimbriae of actinomyces recognize RPS on the streptococci. Cells reactive with antibody against type 2 fimbriae of
Actinomyces naeslundii
T14V (anti-type-2) were much less frequent than either subset of streptococci. However, bacteria reactive with anti-type-2 were seen in intimate association with anti-RPS-reactive cells. These results are the first direct demonstration of coaggregation-mediated interactions during initial plaque accumulation in vivo. Further, these results demonstrate the spatiotemporal development and prevalence of mixed-species communities in early dental plaque.
Role of Feedback Regulation of Pantothenate Kinase (CoaA) in Control of Coenzyme A Levels in
Escherichia coli
Authors: Rock Charles O. , Park Hee-Won , Jackowski Suzanne 
Published: May 2003

ABSTRACT

Pantothenate kinase (CoaA) is a key regulator of coenzyme A (CoA) biosynthesis in
Escherichia coli
, and its activity is controlled by feedback inhibition by CoA and its thioesters. The importance of feedback inhibition in the control of the intracellular CoA levels was tested by constructing three site-directed mutants of CoaA that were predicted to be feedback resistant based on the crystal structure of the CoaA-CoA binary complex. CoaA[R106A], CoaA[H177Q], and CoaA[F247V] were purified and shown to retain significant catalytic activity and be refractory to inhibition by CoA. CoaA[R106A] retained 50% of the catalytic activity of CoaA, whereas the CoaA[H177Q] and CoaA[F247V] mutants were less active. The importance of feedback control of CoaA to the intracellular CoA levels was assessed by expressing either CoaA or CoaA[R106A] in strain ANS3 [
coaA15
(Ts)
panD2
]. Cells expressing CoaA[R106A] had significantly higher levels of phosphorylated pantothenate-derived metabolites and CoA in vivo and excreted significantly more 4′-phosphopantetheine into the medium compared to cells expressing the wild-type protein. These data illustrate the key role of feedback regulation of pantothenate kinase in the control of intracellular CoA levels.