bluuebunny/crossref_metadata_2025_split huggingface dataset:
Fusarium oxysporum
as a Multihost Model for the Genetic Dissection of Fungal Virulence in Plants and Mammals
Authors: Ortoneda Montserrat , Guarro Josep , Madrid Marta P. , Caracuel Zaira , Roncero M. Isabel G. , Mayayo Emilio , Di Pietro Antonio 
Published: February 2004

ABSTRACT

Fungal pathogens cause disease in plant and animal hosts. The extent to which infection mechanisms are conserved between both classes of hosts is unknown. We present a dual plant-animal infection system based on a single strain of
Fusarium oxysporum
, the causal agent of vascular wilt disease in plants and an emerging opportunistic human pathogen. Injection of microconidia of a well-characterized tomato pathogenic isolate (isolate 4287) into the lateral tail vein of immunodepressed mice resulted in disseminated infection of multiple organs and death of the animals. Knockout mutants in genes encoding a mitogen-activated protein kinase, a pH response transcription factor, or a class V chitin synthase previously shown to be implicated in virulence on tomato plants were tested in the mouse model. The results indicate that some of these virulence factors play functionally distinct roles during the infection of tomato plants and mice. Thus, a single
F. oxysporum
strain can be used to study fungal virulence mechanisms in plant and mammalian pathogenesis.
C3 Promotes Clearance ofKlebsiella pneumoniaeby A549 Epithelial Cells
Authors: de Astorza Beatriz , Cortés Guadalupe , Crespí Catalina , Saus Carles , Rojo José María , Albertí Sebastián 
Published: February 2004

ABSTRACTThe airway epithelium represents a primary site for contact between microbes and their hosts. To assess the role of complement in this event, we studied the interaction between the A549 cell line derived from human alveolar epithelial cells and a major nosocomial pathogen,Klebsiella pneumoniae, in the presence of serum. In vitro, we found that C3 opsonization of poorly encapsulatedK. pneumoniaeclinical isolates and an unencapsulated mutant enhanced dramatically bacterial internalization by A549 epithelial cells compared to highly encapsulated clinical isolates. Local complement components (either present in the human bronchoalveolar lavage or produced by A549 epithelial cells) were sufficient to opsonizeK. pneumoniae. CD46 could competitively inhibit the internalization ofK. pneumoniaeby the epithelial cells, suggesting that CD46 is a receptor for the binding of complement-opsonizedK. pneumoniaeto these cells. We observed that poorly encapsulated strains appeared into the alveolar epithelial cells in vivo but that (by contrast) they were completely avirulent in a mouse model of pneumonia compared to the highly encapsulated strains. Our results show that bacterial opsonization by complement enhances the internalization of the avirulent microorganisms by nonphagocytic cells such as A549 epithelial cells and allows an efficient innate defense.
Identification and Molecular Characterization of EatA, an Autotransporter Protein of Enterotoxigenic
Escherichia coli
Authors: Patel Seema K. , Dotson Jimmie , Allen Kenneth P. , Fleckenstein James M. 
Published: February 2004

ABSTRACT

Enterotoxigenic
Escherichia coli
(ETEC) strains remain a formidable cause of diarrheal disease. To identify novel surface proteins of ETEC, we performed Tn
phoA
mutagenesis of prototype ETEC strain H10407 and discovered a secreted protein not previously recognized in ETEC. DNA sequencing of the interrupted locus in mutant Tn
phoA
.977 revealed a candidate 4,095-bp open reading frame without significant homology to commensal
E. coli
K-12 genomic DNA. Translation of this sequence revealed that it encoded a predicted peptide of 147.7 kDa that bears significant homology to members of the autotransporter family of bacterial virulence factors, particularly the serine protease autotransporters of the
Enterobacteriaceae
proteins. The gene identified in H10407,
eatA
(ETEC autotransporter A), encodes a potential serine protease motif (GDSGSP) in the secreted amino-terminal domain, and the predicted peptide shows more than 80% homology with SepA, a virulence protein secreted by
Shigella flexneri
. DNA hybridization and PCR demonstrated that
eatA
resides on the 92-kDa pCS1 virulence plasmid of H10407 and that it is present in multiple clinical ETEC strains. Immunoblots with antisera directed against a recombinant EatA passenger protein fragment identified a 110-kDa protein in supernatants purified from H10407 but not from the Tn
phoA
.977 mutant or H10407-P, which lacks pCS1. EatA possesses serine protease activity that is abolished by mutations within a serine protease catalytic triad formed by residues H
134
, D
162
, and S
267
. Finally, interruption of the
eatA
gene retarded fluid accumulation in the rabbit ileal loop model, suggesting that this autotransporter contributes to the virulence of ETEC.
Inhibitory Effect of Antiserum to Surface Antigen P50 of
Babesia gibsoni
on Growth of Parasites in Severe Combined Immunodeficiency Mice Given Canine Red Blood Cells
Authors: Fukumoto Shinya , Xuan Xuenan , Takabatake Noriyuki , Igarashi Ikuo , Sugimoto Chihiro , Fujisaki Kozo , Nagasawa Hideyuki , Mikami Takeshi , Suzuki Hiroshi 
Published: February 2004

ABSTRACT

The inhibitory effect of an antiserum to surface protein P50 of
Babesia gibsoni
on the growth of the parasite was determined with severe combined immunodeficiency mice given canine red blood cells. The antiserum to the recombinant P50 protein significantly inhibited the parasite growth, indicating that P50 might be a useful vaccine candidate.
Identification of
srv
, a PrfA-Like Regulator of Group A
Streptococcus
That Influences Virulence
Authors: Reid Sean D. , Montgomery Alison G. , Musser James M. 
Published: February 2004

ABSTRACT

We have identified a Crp/Fnr-like transcriptional regulator of
Streptococcus pyogenes
that when inactivated attenuates virulence. The gene, named
srv
for
s
treptococcal regulator of virulence, encodes a 240-amino-acid protein with 53% amino acid similarity to PrfA, a transcriptional activator of virulence in
Listeria monocytogenes
.
Characterization of Cytolethal Distending Toxin Genes and Expression in Shiga Toxin-Producing
Escherichia coli
Strains of Non-O157 Serogroups
Authors: Bielaszewska Martina , Fell Marina , Greune Lilo , Prager Rita , Fruth Angelika , Tschäpe Helmut , Schmidt M. Alexander , Karch Helge 
Published: February 2004

ABSTRACT

We identified cytolethal distending toxin and its gene (
cdt
) in 17 of 340 non-O157 Shiga toxin-producing
Escherichia coli
(STEC) strains (serotypes O73:H18, O91:H21, O113:H21, and O153:H18), all of which were
eae
negative.
cdt
is either chromosomal and homologous to
cdt-V
(serotypes O73:H18, O91:H21, and O113:H21) or plasmidborne and identical to
cdt-III
(serotype O153:H18). Among
eae
-negative STEC,
cdt
was associated with disease (
P
= 0.003).
Inclusion of CpG Adjuvant with Plasmid DNA Coding for NcGRA7 Improves Protection against Congenital Neosporosis
Authors: Jenkins Mark , Parker Carolyn , Tuo Wenbin , Vinyard Bryan , Dubey J. P. 
Published: February 2004

ABSTRACT

The present study showed that incorporation of CpG adjuvant into plasmid DNA coding for NcGRA7 antigen resulted in a twofold increase in the level of protection against congenital transfer of
Neospora caninum
. The level of protection was considerably higher than that observed in pups born from dams immunized with nonrecombinant plasmid.
Heterozygous Toll-Like Receptor 2 Polymorphism Does Not Affect Lipoteichoic Acid-Induced Chemokine and Inflammatory Responses
Authors: von Aulock Sonja , Schröder Nicolas W. J. , Traub Stephanie , Gueinzius Katja , Lorenz Eva , Hartung Thomas , Schumann Ralf R. , Hermann Corinna 
Published: February 2004

ABSTRACT

While transfection of
tlr2
conveyed responsiveness to lipoteichoic acid (LTA), the Arg753Gln polymorphic gene could not. LTA induced a stronger chemokine and anti-inflammatory response than lipopolysaccharides did. Blood from heterozygous polymorphic and wild-type donors reacted uniformly to LTA and
Staphylococcus aureus.
Thus, one functional allele for Toll-like receptor 2 suffices for full cytokine response.
Shiga Toxin 1 Targets Bovine Leukemia Virus-Expressing Cells
Authors: Ferens Witold A. , Grauke Luke J. , Hovde Carolyn J. 
Published: February 2004

ABSTRACT

Direct evidence that
Escherichia coli
Shiga toxin (Stx) acts against bovine leukemia virus (BLV)-expressing cells was obtained. The active A subunit of Stx type 1 (StxA1) targeted a selected population of permeable cells expressing BLV and inhibited BLV replication in a culture of bovine peripheral blood mononuclear cells. Cells were cultured with and without StxA1, and at various times cells expressing BLV were identified by being stained with MW1 monoclonal antibody specific for the BLV protein gp51. Before culture, permeable cells were tagged by uptake of one of the following: acetoxymethyl of 2′,7′-
bis
-(2-carboxyethyl)-5-(and 6)-carboxyfluorescein (BCECF), BCECF conjugated to 70-kDa dextran, or 70-kDa dextran conjugated to fluorescein. The tagged cells costaining with anti-gp51 were selectively eliminated in StxA1-treated cultures. Electron microscopy analysis of purified B lymphocytes showed sharply reduced numbers of BLV particles in StxA1-treated cultures.
Role of Class A Penicillin-Binding Proteins in PBP5-Mediated β-Lactam Resistance inEnterococcus faecalis
Authors: Arbeloa Ana , Segal Heidi , Hugonnet Jean-Emmanuel , Josseaume Nathalie , Dubost Lionnel , Brouard Jean-Paul , Gutmann Laurent , Mengin-Lecreulx Dominique , Arthur Michel 
Published: February 2004

ABSTRACTPeptidoglycan polymerization complexes contain multimodular penicillin-binding proteins (PBP) of classes A and B that associate a conserved C-terminal transpeptidase module to an N-terminal glycosyltransferase or morphogenesis module, respectively. InEnterococcus faecalis, class B PBP5 mediates intrinsic resistance to the cephalosporin class of β-lactam antibiotics, such as ceftriaxone. To identify the glycosyltransferase partner(s) of PBP5, combinations of deletions were introduced in all three class A PBP genes ofE. faecalisJH2-2 (ponA,pbpF, andpbpZ). Among mutants with single or double deletions, only JH2-2 ΔponAΔpbpFwas susceptible to ceftriaxone. Ceftriaxone resistance was restored by heterologous expression ofpbpFfromEnterococcus faeciumbut not bymgtencoding the monofunctional glycosyltransferase ofStaphylococcus aureus. Thus, PBP5 partners essential for peptidoglycan polymerization in the presence of β-lactams formed a subset of the class A PBPs ofE. faecalis, and heterospecific complementation was observed with an ortholog fromE. faecium. Site-directed mutagenesis ofpbpFconfirmed that the catalytic serine residue of the transpeptidase module was not required for resistance. None of the three class A PBP genes was essential for viability, although deletion of the three genes led to an increase in the generation time and to a decrease in peptidoglycan cross-linking. As theE. faecalischromosome does not contain any additional glycosyltransferase-related genes, these observations indicate that glycan chain polymerization in the triple mutant is performed by a novel type of glycosyltransferase. The latter enzyme was not inhibited by moenomycin, since deletion of the three class A PBP genes led to high-level resistance to this glycosyltransferase inhibitor.
Plasmid-Dependent Methylotrophy in Thermotolerant
Bacillus methanolicus
Authors: Brautaset Trygve , Jakobsen Øyvind M. , Flickinger Michael C. , Valla Svein , Ellingsen Trond E. 
Published: February 2004

ABSTRACT

Bacillus methanolicus
can efficiently utilize methanol as a sole carbon source and has an optimum growth temperature of 50°C. With the exception of mannitol, no sugars have been reported to support rapid growth of this organism, which is classified as a restrictive methylotroph. Here we describe the DNA sequence and characterization of a 19,167-bp circular plasmid, designated pBM19, isolated from
B. methanolicus
MGA3. Sequence analysis of pBM19 demonstrated the presence of the methanol dehydrogenase gene,
mdh
, which is crucial for methanol consumption in this bacterium. In addition, five genes (
pfk
, encoding phosphofructokinase;
rpe
, encoding ribulose-5-phosphate 3-epimerase;
tkt
, encoding transketolase;
glpX
, encoding fructose-1,6-bisphosphatase; and
fba
, encoding fructose-1,6-bisphosphate aldolase) with deduced roles in methanol assimilation via the ribulose monophosphate pathway are encoded by pBM19. A shuttle vector, pTB1.9, harboring the pBM19 minimal replicon (
repB
and
ori
) was constructed and used to transform MGA3. Analysis of the resulting recombinant strain demonstrated that it was cured of pBM19 and was not able to grow on methanol. A pTB1.9 derivative harboring the complete
mdh
gene could not restore growth on methanol when it was introduced into the pBM19-cured strain, suggesting that additional pBM19 genes are required for consumption of this carbon source. Screening of 13 thermotolerant
B. methanolicus
wild-type strains showed that they all harbor plasmids similar to pBM19, and this is the first report describing plasmid-linked methylotrophy in any microorganism. Our findings should have an effect on future genetic manipulations of this organism, and they contribute to a new understanding of the biology of methylotrophs.
Identification and Functional Characterization of the
Lactococcus lactis rfb
Operon, Required for dTDP-Rhamnose Biosynthesis
Authors: Boels Ingeborg C. , Beerthuyzen Marke M. , Kosters Marit H. W. , Van Kaauwen Martijn P. W. , Kleerebezem Michiel , de Vos Willem M. 
Published: February 2004

ABSTRACT

dTDP-rhamnose is an important precursor of cell wall polysaccharides and rhamnose-containing exopolysaccharides (EPS) in
Lactococcus lactis
. We cloned the
rfbACBD
operon from
L. lactis
MG1363, which comprises four genes involved in dTDP-rhamnose biosynthesis. When expressed in
Escherichia coli
, the lactococcal
rfbACBD
genes could sustain heterologous production of the
Shigella flexneri
O antigen, providing evidence of their functionality. Overproduction of the RfbAC proteins in
L. lactis
resulted in doubled dTDP-rhamnose levels, indicating that the endogenous RfbAC activities control the intracellular dTDP-rhamnose biosynthesis rate. However, RfbAC overproduction did not affect rhamnose-containing B40-EPS production levels. A nisin-controlled conditional RfbBD mutant was unable to grow in media lacking the inducer nisin, indicating that the
rfb
genes have an essential role in
L. lactis
. Limitation of RfbBD activities resulted in the production of altered EPS. The monomeric sugar of the altered EPS consisted of glucose, galactose, and rhamnose at a molar ratio of 1:0.3:0.2, which is clearly different from the ratio in the native sugar. Biophysical analysis revealed a fourfold-greater molecular mass and a twofold-smaller radius of gyration for the altered EPS, indicating that these EPS are more flexible polymers with changed viscosifying properties. This is the first indication that enzyme activity at the level of central carbohydrate metabolism affects EPS composition.
Capsule Shields the Function of Short Bacterial Adhesins
Authors: Schembri Mark A. , Dalsgaard Dorte , Klemm Per 
Published: February 2004

ABSTRACT

Bacterial surface structures such as capsules and adhesins are generally regarded as important virulence factors. Here we demonstrate that capsules block the function of the self-recognizing protein antigen 43 through physical shielding. The phenomenon is not restricted to
Escherichia coli
but can occur in other gram-negative bacteria. Likewise, we show that other short adhesins exemplified by the AIDA-I protein are blocked by the presence of a capsule. The results support the notion that capsule polysaccharides sterically prevent receptor-target recognition of short bacterial adhesins. This negative interference has important biological consequences, such as affecting the ability of bacteria to form biofilms.
Differential Impact of MexB Mutations on Substrate Selectivity of the MexAB-OprM Multidrug Efflux Pump of
Pseudomonas aeruginosa
Authors: Middlemiss Jocelyn K. , Poole Keith 
Published: February 2004

ABSTRACT

The integral inner membrane resistance-nodulation-division (RND) components of three-component RND-membrane fusion protein-outer membrane factor multidrug efflux systems define the substrate selectivity of these efflux systems. To gain a better understanding of what regions of these proteins are important for substrate recognition, a plasmid-borne
mexB
gene encoding the RND component of the MexAB-OprM multidrug efflux system of
Pseudomonas aeruginosa
was mutagenized in vitro by using hydroxylamine and mutations compromising the MexB contribution to antibiotic resistance identified in a Δ
mexB
strain. Of 100 mutants that expressed wild-type levels of MexB and showed increased susceptibility to one or more of carbenicillin, chloramphenicol, nalidixic acid, and novobiocin, the
mexB
genes of a representative 46 were sequenced, and 19 unique single mutations were identified. While the majority of mutations occurred within the large periplasmic loops between transmembrane segment 1 (TMS-1) and TMS-2 and between TMS-7 and TMS-8 of MexB, mutations were seen in the TMSs and in other periplasmic as well as cytoplasmic loops. By threading the MexB amino acid sequence through the crystal structure of the homologous RND transporter from
Escherichia coli
, AcrB, a three-dimensional model of a MexB trimer was obtained and the mutations were mapped to it. Unexpectedly, most mutations mapped to regions of MexB predicted to be involved in trimerization or interaction with MexA rather than to regions expected to contribute to substrate recognition. Intragenic second-site suppressor mutations that restored the activity of the G220S mutant version of MexB, which was compromised for resistance to all tested MexAB-OprM antimicrobial substrates, were recovered and mapped to the apparently distal portion of MexB that is implicated in OprM interaction. As the G220S mutation likely impacted trimerization, it appears that either proper assembly of the MexB trimer is necessary for OprM interaction or OprM association with an unstable MexB trimer might stabilize it, thereby restoring activity.
The PhoP-PhoQ Two-Component Regulatory System of
Photorhabdus luminescens
Is Essential for Virulence in Insects
Authors: Derzelle Sylviane , Turlin Evelyne , Duchaud Eric , Pages Sylvie , Kunst Frank , Givaudan Alain , Danchin Antoine 
Published: February 2004

ABSTRACT

Photorhabdus luminescens
is a symbiont of entomopathogenic nematodes. Analysis of the genome sequence of this organism revealed a homologue of PhoP-PhoQ, a two-component system associated with virulence in intracellular bacterial pathogens. This organism was shown to respond to the availability of environmental magnesium. A mutant with a knockout mutation in the regulatory component of this system (
phoP
) had no obvious growth defect. It was, however, more motile and more sensitive to antimicrobial peptides than its wild-type parent. Remarkably, the mutation eliminated virulence in an insect model. No insect mortality was observed after injection of a large number of the
phoP
bacteria, while very small amounts of parental cells killed insect larvae in less than 48 h. At the molecular level, the PhoPQ system mediated Mg
2+
-dependent modifications in lipopolysaccharides and controlled a locus (
pbgPE
) required for incorporation of 4-aminoarabinose into lipid A. Mg
2+
-regulated gene expression of
pbgP1
was absent in the mutant and was restored when
phoPQ
was complemented in
trans
. This finding highlights the essential role played by PhoPQ in the virulence of an entomopathogen.
Evidence for a Symbiosis Island Involved in Horizontal Acquisition of Pederin Biosynthetic Capabilities by the Bacterial Symbiont of
Paederus fuscipes
Beetles
Authors: Piel Jörn , Höfer Ivonne , Hui Dequan 
Published: February 2004

ABSTRACT

Pederin belongs to a group of antitumor compounds found in terrestrial beetles and marine sponges. It is used by apparently all members of the rove beetle genera
Paederus
and
Paederidus
as a chemical defense against predators. However, a recent analysis of the putative pederin biosynthesis (
ped
) gene cluster strongly suggests that pederin is produced by bacterial symbionts. We have sequenced an extended region of the symbiont genome to gain further insight into the biology of this as-yet-unculturable bacterium and the evolution of pederin symbiosis. Our data indicate that the symbiont is a very close relative of
Pseudomonas aeruginosa
that has acquired several foreign genetic elements by horizontal gene transfer. Besides one functional tellurite resistance operon, the region contains a genomic island spanning 71.6 kb that harbors the putative pederin biosynthetic genes. Several decayed insertion sequence elements and the mosaic-like appearance of the island suggest that the acquisition of the
ped
symbiosis genes was followed by further insertions and rearrangements. A horizontal transfer of genes for the biosynthesis of protective substances could explain the widespread occurrence of pederin-type compounds in unrelated animals from diverse habitats.
Transcriptional Regulation of Genes Encoding Arabinan-Degrading Enzymes in
Bacillus subtilis
Authors: Raposo Maria Paiva , Inácio José Manuel , Mota Luís Jaime , de Sá-Nogueira Isabel 
Published: February 2004

ABSTRACT

Bacillus subtilis
produces hemicellulases capable of releasing arabinosyl oligomers and arabinose from plant cell walls. In this work, we characterize the transcriptional regulation of three genes encoding arabinan-degrading enzymes that are clustered with genes encoding enzymes that further catabolize arabinose. The
abfA
gene comprised in the metabolic operon
araABDLMNPQ
-
abfA
and the
xsa
gene located 23 kb downstream most probably encode α-
l
-arabinofuranosidases (EC 3.2.1.55). Here, we show that the
abnA
gene, positioned immediately upstream from the metabolic operon, encodes an endo-α-1,5-arabinanase (EC 3.2.1.99). Furthermore, by in vivo RNA studies, we inferred that
abnA
and
xsa
are monocistronic and are transcribed from σ
A
-like promoters. Transcriptional fusion analysis revealed that the expression of the three arabinases is induced by arabinose and arabinan and is repressed by glucose. The levels of induction by arabinose and arabinan are higher during early postexponential growth, suggesting a temporal regulation. Moreover, the induction mechanism of these genes is mediated through negative control by the key regulator of arabinose metabolism, AraR. Thus, we analyzed AraR-DNA interactions by in vitro quantitative DNase I footprinting and in vivo analysis of single-base-pair substitutions within the promoter regions of
xsa
and
abnA
. The results indicate that transcriptional repression of the
abfA
and
xsa
genes is achieved by a tightly controlled mechanism but that the regulation of
abnA
is more flexible. We suggest that the expression of genes encoding extracellular degrading enzymes of arabinose-containing polysaccharides, transport systems, and intracellular enzymes involved in further catabolism is regulated by a coordinate mechanism triggered by arabinose via AraR.