bluuebunny/crossref_metadata_2025 huggingface dataset:
Highlights in the Historical Development of the Porous Media Theory: Toward a Consistent Macroscopic Theory
Authors: de Boer Reint
Published: April 2009

First contributions to the theory of porous media were made by R Woltman in 1794 when he independently developed a more sophisticated earth pressure theory than C Coulomb, and surprisingly in another context introduced the concept of volume fractions. In the last century, further important contributions were published by A Delesse, A Fick, H Darcy, and J Stefan on the concept of surface fractions, the diffusion problem, ground-water flow, and the mixture theory, which are essential parts of the theory of porous media. In the twentieth century, the scientific discussion on porous media theories was opened by P Fillunger in 1913 in a paper about the uplift problem in saturated rigid porous solids. In subsequent articles, he investigated the phenomena of friction and capillarity and discovered the effect of effective stresses. In 1923, K von Terzaghi, founder of modern soil mechanics, started his investigations on saturated deformable porous solids within the framework of the calculation of the permeability coefficient of clay. In 1936, Fillunger founded the concept of the mechancial theory of liquid-saturated deformable porous solids. However, his substantial masterpiece was completely forgotten and ignored. The reason for this may lie in the fact that in the 1930’s deep hostility arose between Fillunger and von Terzaghi due to different scientific views on the porous media theory and soil mechanics, leading to a large controversy which ended very tragically. The works of von Terzaghi and Fillunger were continued by M Biot, G Heinrich, and I Frenkel in the next decades. Today, two important directions of the macroscopic porous media theory are commonly acknowledged. The first one is based on investigations by M Biot, and the second one proceeds from the mixture theory, restricted by the concept of volume fractions (porous media theory). In particular, the porous media theory has turned out to be an efficient tool to treat saturated and empty porous solids.
Modern Developments in Flow Control
Authors: Gad-el-Hak Mohamed
Published: April 2009

This brief article reviews the important advances in the field of flow control that took place during the past few years. This broad area of research remains of great interest for its numerous potential benefits for both the military and civilian sectors. Spurred by the recent developments in chaos control, microfabrication and neural networks, reactive control of turbulent flows is now in the realm of the possible for future practical devices. Other less complex control schemes, passive as well as active, are more market ready and are also witnessing resurgence of interest.
Main Polish Historical and Modern Sources on Applied Mechanics
Authors: Telega J. J. , Wojnar R.
Published: April 2009

The aim of this article is to present the list of books by Polish scientists on applied mechanics in a rather broad sense. An effort was made to cover the last century. Previous Polish sources, until 1874, were collected by Kucharzewski (1894). A list of the main journals in applied mechanics is also given, essentially covering the period starting after the Second World War.
Recent Developments Concerning Saint-Venant’s Principle: A Second Update
Authors: Horgan C. O.
Published: April 2009

Saint-Venant’s principle plays a central role in the theory and applications of elasticity. A comprehensive review of contemporary research on this topic was given by Horgan and Knowles (1983), and updated by Horgan (1989a). Since these articles were written, several developments in the subject area have taken place. In his second update, we review some of this progress, including a discussion of analogous issues arising in entry flow problems for viscous incompressible fluids.
Compliant Coatings: A Decade of Progress
Authors: Gad-el-Hak Mohamed
Published: April 2009

This brief article reviews the important developments in the field of compliant coatings that took place in the past ten years. During this period progress in theoretical and computational methods somewhat outpaced that in experimental efforts. There is no doubt that compliant coatings can be designed to delay transition and to suppress noise on marine vehicles as well as other practical hydrodynamic devices. Transition Reynolds numbers that exceed by an order of magnitude those on rigid-surface boundary layers can be achieved. There is renewed evidence of favorable interactions of compliant coatings even for air flows and even for turbulent boundary layers, but more research is needed to confirm these latest results.
Finite Elements in Aeroelasticity of Plates and Shells
Authors: Bismarck-Nasr Maher N.
Published: April 2009

A review of the finite element method applied to the problem of supersonic aeroelastic stability of nonlinear flat plates, linear, and nonlinear curved plates is presented. Some new contributions in the field are given and future trends are discussed.
Review of Knee Models: 1996 Update
Authors: Hefzy Mohamed Samir , Cooke T. Derek V.
Published: April 2009

This paper is an update on our previous review of knee models (Hefzy and Grood [30]). We find that progress was made in the area of dynamic modeling. Since 1988, a technique was developed to solve the system of differential algebraic equations describing the three-dimensional dynamic behavior of the knee joint. This technique allows to solve complex problems. However, not a single dynamic comprehensive anatomically based mathematical three-dimensional model of the knee joint that includes both tibio-femoral and patello-femoral joints has yet been developed. In the area of meniscal modeling, we find that only one three-dimensional model of the knee joint that includes the menisci is available in the literature. This quasi-static model includes menisci, tibial, femoral and patellar cartilage layers, and ligamentous structures. This model is limited since it solves only for one position of the knee joint: full extension. From this updated survey, we conclude that the development of a comprehensive three-dimensional anatomically based mathematical model of the knee joint (that solves the three-body contact problem at the tibio-femoral joint between the menisci, tibia and femur, and includes the patello-femoral joint) continues to present major challenge.
Acoustic Scattering Resonances: Relation to External and Internal Surface Waves
Authors: U¨berall Herbert , Ge´rard Alain , Guran Arde´shir , Duclos Jean , El Hocine Khelil Mohammed , Bao X. L. , Raju P. K.
Published: April 2009

The resonance scattering theory (RST) and the singularity expansion method (SEM) are both based on the complex-frequency poles of the scattering amplitude in the scattering of acoustic, elastic, or electromagnetic waves from elastic or impenetrable objects, or from cavities. These poles, situated off the real frequency axis at locations with negative imaginary parts, are found to yield, at the real frequencies of the experiments, prominent resonances for acoustic and elastic-wave scattering from elastic objects as discussed in our earlier review (U¨berall et al, Appl Mech Rev43(10), 1990, 235). However, as the authors demonstrated before (U¨berall et al, J Acoust Soc Am61, 1977, 711), the origin of these resonances lies in the phase matching of circumferential or surface waves generated on the target objects during the scattering; hence a study of the resonances will lead to an understanding of, and information on these surface waves. This has been the topic of a large number of studies in recent years, and the results are summarized in the present review for immersed elastic target objects of plane, spherical, and cylindrical geometry, including both elastic-type and fluid-borne surface waves. For multilayered elastic structures, we also describe possible layer-resonance identifications based on acoustic and elastic-wave scattering experiments.
Update of Conventional and Unconventional Plastic Response and Representation
Authors: Drucker Daniel C.
Published: April 2009

The primary purpose of this update is to outline progress that has been made in three directions: 1) exhibiting the real but limited instability of configuration and instability of path for loaded homogeneous specimens of a material model that obeys a non-associated flow rule; 2) exploring the roles of inhomogeneity of material and of stress as explanations for the suitability of idealizing the macroscopic plastic behavior of most structural metals and alloys as stable and time-independent over much of their range of use, despite possible instability on the miniscale and the always-present strong instability and time-dependence on the microscale; and 3) the search for a clearer picture of the likely relation of macroscopic flow stress to properties of dislocation arrays and barrier regions. Other topics, touched upon very briefly, include: finite elastoplasticity, the application of thermodynamics, and the increasing variety of non-traditional or unconventional formulations.
Draft Stem Cell Guidelines Please Many, Disappoint Some
Authors: Holden Constance , Kaiser Jocelyn
Published: April 2009

They are not perfect, but they're a big improvement over what scientists have been living with since 2001. That's how many scientists feel about the draft guidelines on human stem cell research released last week by the U.S. National Institutes of Health.
Renal Disposition of Colistin in the Isolated Perfused Rat Kidney
Authors: Ma Zheng , Wang Jiping , Nation Roger L. , Li Jian , Turnidge John D. , Coulthard Kingsley , Milne Robert W.
Published: April 2009

Nephrotoxicity is an important limitation to the clinical use of colistin againstPseudomonas aeruginosaand other gram-negative pathogens. Previous work reported net tubular reabsorption of colistin by the kidney in vivo, but there is no knowledge of its disposition within the kidney. This study investigated the renal disposition and potential transport mechanisms of colistin in the isolated perfused rat kidney (IPK) model by perfusing with colistin sulfate alone (2 μg/ml) or in the presence of potential inhibitors (tetraethylammonium [TEA], glycine-glycine [Gly-Gly], or hydrochloric acid [HCl]) at three different concentrations. When perfused alone, the renal clearances (CLR) for colistin A and B (the major components of colistin) in control kidneys were constant and low (mean values < 0.05 ml/min throughout the perfusion). The mean clearance ratios [CR, defined as CLR/(fu× GFR), wherefuis the fraction of drug unbound in perfusate and GFR is the glomerular filtration rate] were significantly less than 1. It was concluded that there is net tubular reabsorption of colistin, and this exceeded the reabsorption of water. Less than 10% eliminated from perfusate was recovered in urine, suggesting considerable renal accumulation of colistin. The CR values for colistin were significantly increased when perfused with TEA (500 μM), Gly-Gly (833 μM), and HCl (2,500, 5,000, and 10,000 μM). It is proposed that renal reabsorption of colistin may involve organic cation transporters (inhibited by TEA) and peptide transporters (inhibited by Gly-Gly) and that the process is sensitive to the pH of urine.
First Detection of Plasmid-Encoded
bla
OXY
β-Lactamase
Authors: González-López J. J. , Coelho A. , Larrosa M. N. , Lavilla S. , Bartolomé R. , Prats G.
Published: April 2009

Three
Klebsiella oxytoca
isolates and one
Klebsiella pneumoniae
isolate from three children admitted to the Hematology Unit of Hospital Vall d'Hebron (Barcelona, Spain) exhibited a susceptibility pattern suggesting OXY β-lactamase hyperproduction. All the isolates contained a 95-kb plasmid that harbored
bla
OXY-1
, which was transferred by electrotransformation but could not be self-transferred by conjugation. A
qnrS1
gene was also harbored in the
bla
OXY-1
-carrying plasmid. This is the first report of a plasmid-encoded OXY β-lactamase.
Prolonged Impact of Antibiotics on Intestinal Microbial Ecology and Susceptibility to Enteric
Salmonella
Infection
Authors: Croswell Amy , Amir Elad , Teggatz Paul , Barman Melissa , Salzman Nita H.
Published: April 2009

The impact of antibiotics on the host's protective microbiota and the resulting increased susceptibility to mucosal infection are poorly understood. In this study, antibiotic regimens commonly applied to murine enteritis models are used to examine the impact of antibiotics on the intestinal microbiota, the time course of recovery of the biota, and the resulting susceptibility to enteric
Salmonella
infection. Molecular analysis of the microbiota showed that antibiotic treatment has an impact on the colonization of the murine gut that is site and antibiotic dependent. While combinations of antibiotics were able to eliminate culturable bacteria, none of the antibiotic treatments were effective at sterilizing the intestinal tract. Recovery of total bacterial numbers occurs within 1 week after antibiotic withdrawal, but alterations in specific bacterial groups persist for several weeks. Increased
Salmonella
translocation associated with antibiotic pretreatment corrects rapidly in association with the recovery of the most dominant bacterial group, which parallels the recovery of total bacterial numbers. However, susceptibility to intestinal colonization and mucosal inflammation persists when mice are infected several weeks after withdrawal of antibiotics, correlating with subtle alterations in the intestinal microbiome involving alterations of specific bacterial groups. These results show that the colonizing microbiotas are integral to mucosal host protection, that specific features of the microbiome impact different aspects of enteric
Salmonella
pathogenesis, and that antibiotics can have prolonged deleterious effects on intestinal colonization resistance.
Shiga Toxin 2 Is Specifically Released from Bacterial Cells by Two Different Mechanisms
Authors: Shimizu Takeshi , Ohta Yuko , Noda Masatoshi
Published: April 2009

Shiga toxin 1 (Stx1) is located in the periplasmic fraction, while Stx2 is found in the extracellular fraction, suggesting that enterohemorrhagic
Escherichia coli
(EHEC) contains a specific Stx2 release system. Both
stx
1
and
stx
2
are found within the late operons of Stx-encoding phages. Stx2 production is greatly induced by mitomycin C, suggesting that
stx
2
can transcribe from the late phage promoter of the Stx2-encoding phage. However, the Stx1 promoter adjacent to
stx
1
is a dominant regulatory element in Stx1 production. With the deletion of phage lysis genes of the Stx2-encoding phage, Stx2 remains in the bacterial cells. Further, we demonstrate that the Stx2-encoding phage, but not the Stx1-encoding phage, is spontaneously induced at extremely low rates. These results indicate that spontaneously specific Stx2-encoding phage induction is involved in specific Stx2 release from bacterial cells. Furthermore, to examine whether another system for specific Stx2 release is present in EHEC, we analyze the
stx
-replaced mutants. As expected, Stx2 derived from the Stx1 promoter is located in both the extracellular and cell-associated fractions, while mutant Stx2 (B subunit, S31N) derived from the Stx1 promoter is found only in the cell-associated fraction. These results indicate that EHEC has another Stx2 release system that strictly recognizes the serine 31 residue of the B subunit. Overall, we present evidence that specific Stx2 release from bacterial cells is involved in both the Stx2-encoding phage induction system and another Stx2 release system.
Cellular Tumor Necrosis Factor, Gamma Interferon, and Interleukin-6 Responses as Correlates of Immunity and Risk of Clinical
Plasmodium falciparum
Malaria in Children from Papua New Guinea
Authors: Robinson Leanne J. , D'Ombrain Marthe C. , Stanisic Danielle I. , Taraika Jack , Bernard Nicholas , Richards Jack S. , Beeson James G. , Tavul Livingstone , Michon Pascal , Mueller Ivo , Schofield Louis
Published: April 2009

The role of early to intermediate
Plasmodium falciparum
-induced cellular responses in the development of clinical immunity to malaria is not well understood, and such responses have been proposed to contribute to both immunity and risk of clinical malaria episodes. To investigate whether
P. falciparum
-induced cellular responses are able to function as predictive correlates of parasitological and clinical outcomes, we conducted a prospective cohort study of children (5 to 14 years of age) residing in a region of Papua New Guinea where malaria is endemic Live, intact
P. falciparum
-infected red blood cells were applied to isolated peripheral blood mononuclear cells obtained at baseline. Cellular cytokine production, including production of interleukin-2 (IL-2), IL-4, IL-6, IL-10, tumor necrosis factor (TNF) (formerly tumor necrosis factor alpha), and gamma interferon (IFN-γ), was measured, and the cellular source of key cytokines was investigated. Multicytokine models revealed that increasing
P. falciparum
-induced IL-6 production was associated with an increased incidence of
P. falciparum
clinical episodes (incidence rate ratio [IRR], 1.75; 95% confidence interval [CI], 1.20 to 2.53), while increasing
P. falciparum
-induced TNF and IFN-γ production was associated with a reduced incidence of clinical episodes (IRR for TNF, 0.55 [95% CI, 0.38 to 0.80]; IRR for IFN-γ, 0.71 [95% CI, 0.55 to 0.90]). Furthermore, we found that monocytes/macrophages and γδ-T cells are important for the
P. falciparum
-induced production of IL-6 and TNF. Early to intermediate cellular cytokine responses to
P. falciparum
may therefore be important correlates of immunity and risk of symptomatic malaria episodes and thus warrant detailed investigation in relation to the development and implementation of effective vaccines.
The Arginine Catabolic Mobile Element Is Not Associated with Enhanced Virulence in Experimental Invasive Disease Caused by the Community-Associated Methicillin-Resistant
Staphylococcus aureus
USA300 Genetic Background
Authors: Montgomery Christopher P. , Boyle-Vavra Susan , Daum Robert S.
Published: April 2009

USA300 has become the predominant community-associated methicillin (meticillin)-resistant
Staphylococcus aureus
(CA-MRSA) genetic background in most U.S. communities. The reasons for the dominance of this genetic background are unclear, but the presence of the recently identified arginine catabolic mobile element (ACME) in the USA300 genome has been advocated as one possibility. CA-MRSA clinical isolates (USA300) differing in the presence or absence of ACME and a USA300 wild-type/ACME deletion mutant pair were analyzed for in vitro expression of global regulatory genes and production of virulence factors. The virulence of these isolates was compared in rodent models of necrotizing pneumonia and skin infection. There was no significant difference in the expression of selected genes mediating virulence (
hla
,
lukSF-PV
,
agr
,
saeRS
) among the isolates tested, regardless of the presence of ACME. There was a higher abundance of α-hemolysin in culture supernatants among ACME-positive isolates than among ACME-negative isolates, but there was no significant difference in the levels of protein A. The presence of ACME was not associated with increased virulence in a rat model of necrotizing pneumonia, as assessed by mortality, in vivo bacterial survival, and severity of lung pathology. Nor was the presence of ACME associated with increased dermonecrosis in a model of skin infection. We conclude that ACME is not necessary for virulence in rodent models of CA-MRSA USA300 pneumonia or skin infection.
Processing of
Pseudomonas aeruginosa
Exotoxin A Is Dispensable for Cell Intoxication
Authors: Morlon-Guyot Juliette , Méré Jocelyn , Bonhoure Anne , Beaumelle Bruno
Published: April 2009

Exotoxin A is a major virulence factor of
Pseudomonas aeruginosa
. This toxin binds to a specific receptor on animal cells, allowing endocytosis of the toxin. Once in endosomes, the exotoxin can be processed by furin to generate a C-terminal toxin fragment that lacks the receptor binding domain and is retrogradely transported to the endoplasmic reticulum for retrotranslocation to the cytosol through the Sec61 channel. The toxin then blocks protein synthesis by ADP ribosylation of elongation factor 2, thereby triggering cell death. A shorter intracellular route has also been described for this toxin. It involves direct translocation of the entire toxin from endosomes to the cytosol and therefore does not rely on furin-mediated cleavage. To examine the implications of endosomal translocation in the intoxication process, we investigated whether the toxin required furin-mediated processing in order to kill cells. We used three different approaches. We first fused to the N terminus of the toxin proteins with different unfolding abilities so that they inhibited or did not inhibit endosomal translocation of the chimera. We then assayed the amount of toxin fragments delivered to the cytosol during cell intoxication. Finally we used furin inhibitors and examined the fate and intracellular localization of the toxin and its receptor. The results showed that exotoxin cytotoxicity results largely from endosomal translocation of the entire toxin. We found that the C-terminal fragment was unstable in the cytosol.
Effects of Amino Acid Substitutions at Conserved and Acidic Residues within Region 1.1 of
Escherichia coli
ς
70
Authors: Bowers Christina Wilson , McCracken Andrea , Dombroski Alicia J.
Published: April 2009

Amino acid substitutions in
Escherichia coli
ς
70
were generated and characterized in an analysis of the role of region 1.1 in transcription initiation. Several acidic and conserved residues are tolerant of substitution. However, replacement of aspartic acid 61 with alanine results in inactivity caused by structural and functional thermolability.
Synthesis of Triacylglycerols by the Acyl-Coenzyme A:Diacyl-Glycerol Acyltransferase Dga1p in Lipid Particles of the Yeast
Saccharomyces cerevisiae
Authors: Sorger Daniel , Daum Günther
Published: April 2009

The terminal step of triacylglycerol (TAG) formation in the yeast
Saccharomyces cerevisiae
is catalyzed by the enzyme acyl-CoA:diacylglycerol acyltransferase (DAGAT). In this study we demonstrate that the gene product of
YOR245c
, Dga1p, catalyzes a major yeast DAGAT activity which is localized to lipid particles. Enzyme measurements employing a newly established assay containing radioactively labeled diacylglycerol (DAG) as a substrate and unlabeled palmitoyl-CoA as a cosubstrate revealed a 70- to 90-fold enrichment of DAGAT in lipid particles over the homogenate but also a 2- to 3-fold enrichment in endoplasmic reticulum fractions. In a
dga1
deletion strain, the DAGAT activity in lipid particles is dramatically reduced, whereas the activity in microsomes is affected only to a minor extent. Thus, we propose the existence of DAGAT isoenzymes in the microsomal fraction. Furthermore, we unveiled an acyl-CoA-independent TAG synthase activity in lipid particles which is distinct from Dga1p and the phosphatidylcholine:DAGAT Lro1p. This acyl-CoA-independent TAG synthase utilizes DAG as an acceptor and free fatty acids as cosubstrates and occurs independently of the acyl-CoA synthases Faa1p to Faa4p. Based on lipid analysis of the respective deletion strains, Lro1p and Dga1p are the major contributors to total cellular TAG synthesis, whereas other TAG synthesizing systems appear to be of minor importance. In conclusion, at least three different pathways are involved in the formation of storage TAG in the yeast.
Effects of Lipoprotein Biogenesis Mutations on Flagellar Assembly in
Salmonella
Authors: Dailey Frank E. , Macnab Robert M.
Published: April 2009

Flagellar assembly requires the expression of a large number of flagellum-specific genes. However, mutations in a number of other genes in
Salmonella
and
Escherichia coli
have been shown to have pleiotropic effects that affect flagellar assembly. FlgH (the L-ring subunit of the flagellar basal body) is a lipoprotein whose modification is important for L-ring assembly. We therefore tested whether the lack of motility of
Salmonella
mutants defective in lipoprotein biogenesis is a result of inability to modify FlgH. Our results show that temperature-sensitive apolipoprotein
N
-acyltransferase [
lnt
(Ts)] mutants are nonflagellate at 42°C. However, the flagellar assembly defect occurs at a much earlier step in the pathway than L-ring assembly. These mutants failed to assemble even an MS ring, presumably because of the observed decrease in transcription of
fliF.
In contrast, temperature-sensitive diacylglycerol transferase [
lgt
(Ts)] mutants were motile at 42°C, provided the strains carried an
lpp
(Braun lipoprotein) mutation to permit growth. We have isolated second-site mutants from an
lgt
(Ts)
lpp
+
strain that grow but are nonflagellate at 42°C. Thus, lipoprotein biogenesis is a factor that is important for flagellar assembly.
Molecular Characterization and Transcriptional Analysis of
adhE2
, the Gene Encoding the NADH-Dependent Aldehyde/Alcohol Dehydrogenase Responsible for Butanol Production in Alcohologenic Cultures of
Clostridium acetobutylicum
ATCC 824
Authors: Fontaine Lisa , Meynial-Salles Isabelle , Girbal Laurence , Yang Xinghong , Croux Christian , Soucaille Philippe
Published: April 2009

The
adhE2
gene of
Clostridium acetobutylicum
ATCC 824, coding for an aldehyde/alcohol dehydrogenase (AADH), was characterized from molecular and biochemical points of view. The 2,577-bp
adhE2
codes for a 94.4-kDa protein.
adhE2
is expressed, as a monocistronic operon, in alcohologenic cultures and not in solventogenic cultures. Primer extension analysis identified two transcriptional start sites 160 and 215 bp upstream of the
adhE2
start codon. The expression of
adhE2
from a plasmid in the DG1 mutant of
C. acetobutylicum
, a mutant cured of the pSOL1 megaplasmid, restored butanol production and provided elevated activities of NADH-dependent butyraldehyde and butanol dehydrogenases. The recombinant AdhE2 protein expressed in
E. coli
as a
Strep
-tag fusion protein and purified to homogeneity also demonstrated NADH-dependent butyraldehyde and butanol dehydrogenase activities. This is the second AADH identified in
C. acetobutylicum
ATCC 824, and to our knowledge this is the first example of a bacterium with two AADHs. It is noteworthy that the two corresponding genes,
adhE
and
adhE2
, are carried by the pSOL1 megaplasmid of
C. acetobutylicum
ATCC 824.
NolX of
Sinorhizobium fredii
USDA257, a Type III-Secreted Protein Involved in Host Range Determination, Is Localized in the Infection Threads of Cowpea (
Vigna unguiculata
[L.] Walp) and Soybean (
Glycine max
[L.] Merr.) Nodules
Authors: Krishnan Hari B.
Published: April 2009

Sinorhizobium fredii
USDA257 forms nitrogen-fixing nodules on soybean (
Glycine max
[L.] Merr.) in a cultivar-specific manner. This strain forms nodules on primitive soybean cultivars but fails to nodulate agronomically improved North American cultivars. Soybean cultivar specificity is regulated by the
nolXWBTUV
locus, which encodes part of a type III secretion system (TTSS). NolX, a soybean cultivar specificity protein, is secreted by TTSS and shows homology to HrpF of the plant pathogen
Xanthomonas campestris
pv. vesicatoria. It is not known whether NolX functions at the bacterium-plant interface or acts inside the host cell. Antibodies raised against
S. fredii
USDA257 NolX were used in immunocytochemical studies to investigate the subcellular localization of this protein. Immunostaining of paraffin-embedded sections of developing soybean and cowpea (
Vigna unguiculata
[L.] Walp) nodules revealed localization of NolX in the infection threads. Protein A-gold immunocytochemical localization studies utilizing affinity-purified NolX antibodies revealed specific deposition of gold particles in the fibrillar material inside infection threads. Similar immunogold localization studies failed to detect NolX in thin sections of mature soybean and cowpea nodules. The results from this study indicate that NolX is expressed in planta only during the early stages of nodule development.
Are Putative Periodontal Pathogens Reliable Diagnostic Markers?
Authors: Riep Birgit , Edesi-Neuß Lilian , Claessen Friderike , Skarabis Horst , Ehmke Benjamin , Flemmig Thomas F. , Bernimoulin Jean-Pierre , Göbel Ulf B. , Moter Annette
Published: April 2009

Periodontitis is one of the most common chronic inflammatory diseases. A number of putative bacterial pathogens have been associated with the disease and are used as diagnostic markers. In the present study, we compared the prevalence of oral bacterial species in the subgingival biofilm of generalized aggressive periodontitis (GAP) (
n
= 44) and chronic periodontitis (CP) (
n
= 46) patients with that of a periodontitis-resistant control group (PR) (
n
= 21). The control group consisted of subjects at least 65 years of age with only minimal or no periodontitis and no history of periodontal treatment. A total of 555 samples from 111 subjects were included in this study. The samples were analyzed by PCR of 16S rRNA gene fragments and subsequent dot blot hybridization using oligonucleotide probes specific for
Aggregatibacter
(
Actinobacillus) actinomycetemcomitans
,
Porphyromonas gingivalis
,
Prevotella intermedia
,
Tannerella forsythia
, a
Treponema denticola
-like phylogroup (Treponema phylogroup II),
Treponema lecithinolyticum
,
Campylobacter rectus
,
Fusobacterium
spp., and
Fusobacterium nucleatum
, as well as
Capnocytophaga ochracea
. Our data confirm a high prevalence of the putative periodontal pathogens
P. gingivalis
,
P. intermedia
, and
T. forsythia
in the periodontitis groups. However, these species were also frequently detected in the PR group. For most of the species tested, the prevalence was more associated with increased probing depth than with the subject group.
T. lecithinolyticum
was the only periodontopathogenic species showing significant differences both between GAP and CP patients and between GAP patients and PR subjects.
C. ochracea
was associated with the PR subjects, regardless of the probing depth. These results indicate that
T. lecithinolyticum
may be a diagnostic marker for GAP and
C. ochracea
for periodontal health. They also suggest that current presumptions of the association of specific bacteria with periodontal health and disease require further evaluation.
Phenotypic and Molecular Characterization of Clinical Isolates of
Mycobacterium elephantis
from Human Specimens
Authors: Turenne Christine , Chedore Pamela , Wolfe Joyce , Jamieson Frances , May Kevin , Kabani Amin
Published: April 2009

Eleven strains of a rapidly growing mycobacterium were isolated from patient specimens originating from various regions of the province of Ontario, Canada, over a 2-year period. Unique high-performance liquid chromatography (HPLC) and PCR-restriction enzyme pattern analysis (PRA) profiles initially suggested a new
Mycobacterium
species, while sequencing of the 16S rRNA gene revealed a sequence match with
Mycobacterium
sp. strain MCRO 17 (GenBank accession no.
X93028
), an isolate determined to be unique which is to date uncharacterized, and also a close similarity to
M
.
elephantis
(GenBank accession no.
AJ010747
), with six base pair variations. A complete biochemical profile of these isolates revealed a species of mycobacteria with phenotypic characteristics similar to those of
M
.
flavescens
. HPLC, PRA, and 16S rRNA sequencing of strain
M
.
elephantis
DSM 44368
T
and result comparisons with the clinical isolates revealed that these strains were in fact
M
.
elephantis
, a newly described species isolated from an elephant. All strains were isolated from human samples, 10 from sputum and 1 from an axillary lymph node.
Development of Infectious Clones for Virulent and Avirulent Pichinde Viruses: a Model Virus To Study Arenavirus-Induced Hemorrhagic Fevers
Authors: Lan Shuiyun , McLay Schelde Lisa , Wang Jialong , Kumar Naveen , Ly Hinh , Liang Yuying
Published: April 2009

Several arenaviruses can cause hemorrhagic fever diseases (VHFs) in humans, the pathogenic mechanism of which is poorly understood due to their virulent nature and the lack of molecular clones. A safe, convenient, and economical small animal model of arenavirus hemorrhagic fever is based on guinea pigs infected by the arenavirus Pichinde (PICV). PICV does not cause disease in humans, but an adapted strain of PICV (P18) causes a disease in guinea pigs that mimics arenavirus hemorrhagic fever in humans in many aspects, while a low-passaged strain (P2) remains avirulent in infected animals. In order to identify the virulence determinants within the PICV genome, we developed the molecular clones for both the avirulent P2 and virulent P18 viruses. Recombinant viruses were generated by transfecting plasmids that contain the antigenomic L and S RNA segments of PICV under the control of the T7 promoter into BSRT7-5 cells, which constitutively express T7 RNA polymerase. By analyzing viral growth kinetics in vitro and virulence in vivo, we show that the recombinant viruses accurately recapitulate the replication and virulence natures of their respective parental viruses. Both parental and recombinant virulent viruses led to high levels of viremia and titers in different organs of the infected animals, whereas the avirulent viruses were effectively controlled and cleared by the hosts. These novel infectious clones for the PICV provide essential tools to identify the virulence factors that are responsible for the severe VHF-like disease in infected animals.
The Respiratory Syncytial Virus M2-1 Protein Forms Tetramers and Interacts with RNA and P in a Competitive Manner
Authors: Tran Thi-Lan , Castagné Nathalie , Dubosclard Virginie , Noinville Sylvie , Koch Emmanuelle , Moudjou Mohammed , Henry Céline , Bernard Julie , Yeo Robert Paul , Eléouët Jean-François
Published: April 2009

The respiratory syncytial virus (RSV) M2-1 protein is an essential cofactor of the viral RNA polymerase complex and functions as a transcriptional processivity and antitermination factor. M2-1, which exists in a phosphorylated or unphosphorylated form in infected cells, is an RNA-binding protein that also interacts with some of the other components of the viral polymerase complex. It contains a CCCH motif, a putative zinc-binding domain that is essential for M2-1 function, at the N terminus. To gain insight into its structural organization, M2-1 was produced as a recombinant protein in
Escherichia coli
and purified to >95% homogeneity by using a glutathione
S
-transferase (GST) tag. The GST-M2-1 fusion proteins were copurified with bacterial RNA, which could be eliminated by a high-salt wash. Circular dichroism analysis showed that M2-1 is largely α-helical. Chemical cross-linking, dynamic light scattering, sedimentation velocity, and electron microscopy analyses led to the conclusion that M2-1 forms a 5.4S tetramer of 89 kDa and ∼7.6 nm in diameter at micromolar concentrations. By using a series of deletion mutants, the oligomerization domain of M2-1 was mapped to a putative α-helix consisting of amino acid residues 32 to 63. When tested in an RSV minigenome replicon system using a luciferase gene as a reporter, an M2-1 deletion mutant lacking this region showed a significant reduction in RNA transcription compared to wild-type M2-1, indicating that M2-1 oligomerization is essential for the activity of the protein. We also show that the region encompassing amino acid residues 59 to 178 binds to P and RNA in a competitive manner that is independent of the phosphorylation status of M2-1.
Lymphocyte and Antibody Responses Reduce Enterovirus 71 Lethality in Mice by Decreasing Tissue Viral Loads
Authors: Lin Yu-Wen , Chang Kung-Chao , Kao Chia-Min , Chang Shih-Ping , Tung Yuk-Ying , Chen Shun-Hua
Published: April 2009

Enterovirus 71 (EV71) infects the central nervous system and causes death and long-term neurological sequelae in hundreds of thousands of young children, but its pathogenesis remains elusive. Immunopathological mechanisms have been suspected to contribute to the pathogenesis of neurological symptoms, so anti-inflammatory agents have been used to treat patients with neurological symptoms. The present study was therefore designed to investigate the functions of lymphocyte and antibody responses in EV71 infection using a mouse model. Immunohistochemical staining analysis revealed virus and three types of lymphocytes, B cells, CD4 T cells, and CD8 T cells, in the spinal cord of an EV71-infected patient who died. A study of mice showed that the levels of virus and lymphocytes in brains and antibody titers in sera were elevated during the time when the mice succumbed to death in a phenomenon analogous to that observed in patients. Further studies demonstrated that after infection, the disease severity, mortality, and tissue viral loads of mice deficient in B, CD4 T, or CD8 T cells were significantly higher than those of wild-type mice. In addition, treatment with a virus-specific antibody, but not a control antibody, before or after infection significantly reduced the disease severity, mortality, and tissue viral loads of mice deficient in B cells. Our results show that both lymphocyte and antibody responses protect mice from EV71 infection. Our study suggests the use of vaccines and virus-specific antibodies to control fatal outbreaks and raises caution over the use of corticosteroids to treat EV71-infected patients with neurological symptoms.
Human Parainfluenza Virus Infection of the Airway Epithelium: Viral Hemagglutinin-Neuraminidase Regulates Fusion Protein Activation and Modulates Infectivity
Authors: Palermo Laura M. , Porotto Matteo , Yokoyama Christine C. , Palmer Samantha G. , Mungall Bruce A. , Greengard Olga , Niewiesk Stefan , Moscona Anne
Published: April 2009

Three discrete activities of the paramyxovirus hemagglutinin-neuraminidase (HN) protein, receptor binding, receptor cleaving (neuraminidase), and triggering of the fusion protein, each affect the promotion of viral fusion and entry. For human parainfluenza virus type 3 (HPIV3), the effects of specific mutations that alter these functions of the receptor-binding protein have been well characterized using cultured monolayer cells, which have identified steps that are potentially relevant to pathogenesis. In the present study, proposed mechanisms that are relevant to pathogenesis were tested in natural host cell cultures, a model of the human airway epithelium (HAE) in which primary HAE cells are cultured at an air-liquid interface and retain functional properties. Infection of HAE cells with wild-type HPIV3 and variant viruses closely reflects that seen in an animal model, the cotton rat, suggesting that HAE cells provide an ideal system for assessing the interplay of host cell and viral factors in pathogenesis and for screening for inhibitory molecules that would be effective in vivo. Both HN′s receptor avidity and the function and timing of F activation by HN require a critical balance for the establishment of ongoing infection in the HAE, and these HN functions independently modulate the production of active virions. Alterations in HN′s F-triggering function lead to the release of noninfectious viral particles and a failure of the virus to spread. The finding that the dysregulation of F triggering prohibits successful infection in HAE cells suggests that antiviral strategies targeted to HN′s F-triggering activity may have promise in vivo.
Activated Ras/MEK Inhibits the Antiviral Response of Alpha Interferon by Reducing STAT2 Levels
Authors: Christian Sherri L. , Collier Thaddeus W. , Zu Dong , Licursi Maria , Hough Chris M. , Hirasawa Kensuke
Published: April 2009

The ability of interferon (IFN) to induce the expression of antiviral genes, and therefore suppress viral infection, is dependent on the activity of cellular suppressors. The Ras/MEK pathway is one of these cellular suppressors, since the activation of Ras/MEK permits viral replication in the presence of alpha IFN (IFN-α). Here, we have investigated the mechanism by which activated Ras/MEK inhibits the IFN-α response. We found that the induction of antiviral proteins in response to IFN-α was impaired in Ras-transformed NIH 3T3 (RasV12) cells. The inhibition of the Ras/MEK pathway restored the IFN-mediated induction of antiviral genes, indicating that activated Ras interrupts the IFN pathway upstream of antiviral gene transcription. Indeed, the IFN-induced phosphorylation of signal transducer and activator of transcription 1 (STAT1) and STAT2 was inhibited in RasV12 cells compared to that of vector control cells. In addition, we found that the total amount of STAT2 was reduced in RasV12 cells. To determine if the impaired IFN-α response can be rescued by restoring the overall level of STAT2, we overexpressed STAT2 in RasV12 cells. The IFN-α-induced phosphorylation of STAT1 and STAT2, as well as the expression of antiviral protein, were restored, and IFN-induced antiviral protection was partially restored. Moreover, we demonstrated that the downregulation of STAT2 levels by Ras/MEK was mediated at the transcriptional level. Thus, the activation of the Ras/MEK pathway reduces the amount of STAT2 available for propagating the IFN signal, resulting in the impairment of the IFN-α-induced antiviral response.
Viral Entry through CXCR4 Is a Pathogenic Factor and Therapeutic Target in Human Immunodeficiency Virus Type 1 Disease
Authors: Schramm Birgit , Penn Michael L. , Speck Roberto F. , Chan Stephen Y. , De Clercq Erik , Schols Dominique , Connor Ruth I. , Goldsmith Mark A.
Published: April 2009

The chemokine receptors CCR5 and CXCR4 function as the principal coreceptors for human immunodeficiency virus type 1 (HIV-1). Coreceptor function has also been demonstrated for a variety of related receptors in vitro. The relative contributions of CCR5, CXCR4, and other putative coreceptors to HIV-1 disease in vivo have yet to be defined. In this study, we used sequential primary isolates and recombinant strains of HIV-1 to demonstrate that CXCR4-using (X4) viruses emerging in association with disease progression are highly pathogenic in ex vivo lymphoid tissues compared to CXCR4-independent viruses. Furthermore, synthetic receptor antagonists that specifically block CXCR4-mediated entry dramatically suppressed the depletion of CD4
+
T cells by recombinant and clinically derived X4 HIV-1 isolates. Moreover, in vitro specificity for the additional coreceptors CCR3, CCR8, BOB, and Bonzo did not augment cytopathicity or diminish sensitivity toward CXCR4 antagonists in lymphoid tissues. These data provide strong evidence to support the concept that adaptation to CXCR4 specificity in vivo accelerates HIV-1 disease progression. Thus, therapeutic intervention targeting the interaction of HIV-1 gp120 with CXCR4 may be highly valuable for suppressing the pathogenic effects of late-stage viruses.
E1B 55-Kilodalton Oncoproteins of Adenovirus Types 5 and 12 Inactivate and Relocalize p53, but Not p51 or p73, and Cooperate with E4orf6 Proteins To Destabilize p53
Authors: Wienzek Sandra , Roth Judith , Dobbelstein Matthias
Published: April 2009

The p53 tumor suppressor protein represents a target for viral and cellular oncoproteins, including adenovirus gene products. Recently, it was discovered that several proteins with structural and functional homologies to p53 exist in human cells. Two of them were termed p51 and p73. We have shown previously that the E1B 55-kDa protein (E1B-55 kDa) of adenovirus type 5 (Ad5) binds and inactivates p53 but not p73. Further, p53 is rapidly degraded in the presence of E1B-55 kDa and the E4orf6 protein of this virus. Here, it is demonstrated that p51 does not detectably associate with E1B-55 kDa. While p53 is relocalized to the cytoplasm by E1B-55 kDa, p51's location is unaffected. Finally, p51 retains its full transcriptional activity in the presence of E1B-55 kDa. Apparently, p51 does not represent a target of Ad5 E1B-55 kDa, suggesting that the functions of p51 are distinct from p53-like tumor suppression. E1B-55 kDa from highly oncogenic adenovirus type 12 (Ad12) was previously shown to surpass the oncogenic activity of Ad5 E1B-55 kDa in various assay systems, raising the possibility that Ad12 E1B-55 kDa might target a broader range of p53-like proteins. However, we show here that Ad12 E1B-55 kDa also inhibits p53's transcriptional activity without measurably affecting p73 or p51. Moderate inhibition of p51's transcriptional activity was observed in the presence of the E4orf6 proteins from Ad5 and Ad12. p53 and Ad12-E1B-55 kDa colocalize in the nucleus and also in cytoplasmic clusters when transiently coexpressed. Finally, E1B-55 kDa and E4orf6 of Ad12 mediate rapid degradation of p53 with an efficiency comparable to that of the Ad5 proteins in human and rodent cells. Our results suggest that E1B-55 kDa of either virus type has similar effects on p53 but does not affect p73 and p51.
Initial Events in Infectious Salmon Anemia Virus Infection: Evidence for the Requirement of a Low-pH Step
Authors: Eliassen Trygve Meum , Frøystad Marianne Kristin , Dannevig Birgit Helene , Jankowska Monika , Brech Andreas , Falk Knut , Romøren Kristine , Gjøen Tor
Published: April 2009

We have investigated the initial steps in the interaction between infectious salmon anemia virus (ISAV) and cultured cells from Atlantic salmon (SHK-1 cell line). Using radioactively or fluorescently labelled viral particles we have studied the binding and fusion kinetics and the effect of pH on binding, uptake, and fusion of ISAV to SHK-1 cells and liposomes. As pH in the medium was reduced from 7.5 to 4.5, the association of virus to the cells was nearly doubled. The same effect of pH was observed when fusion between ISAV and liposomes was analyzed. In addition, the binding of ISAV to intact SHK-1 cells and to cell membrane proteins blotted onto filters was neuraminidase sensitive. However, the increased binding induced by low pH was not neuraminidase sensitive, probably reflecting activation of a fusion peptide at low pH. By using confocal fluorescence microscopy, the increased fusion of fluorescently labelled ISAV with the plasma membrane due to low pH could be demonstrated. When vacuolar pH in the cells was raised during inoculation with chloroquine or ammonium chloride, both electron and confocal microscopy showed accumulation of ISAV in endosomes and lysosomes. Production of infectious virus could be increased by lowering the extracellular pH during infection. Furthermore, chloroquine present during virus inoculation also caused a reduction in the synthesis of viral proteins in ISAV-infected cells as well as in the production of infective virus. These results indicate that ISAV binds to sialic acid residues on the cell surface and that the fusion between virus and cell membrane takes place in the acid environment of endosomes. This provides further evidence for a high degree of similarity between ISAV and influenza virus and extends the basis for the classification of this virus as a member of the
Orthomyxoviridae
family.
Measles Virus Induces Functional TRAIL Production by Human Dendritic Cells
Authors: Vidalain Pierre-Olivier , Azocar Olga , Lamouille Barbara , Astier Anne , Rabourdin-Combe Chantal , Servet-Delprat Christine
Published: April 2009

Measles virus infection induces a profound immunosuppression that can lead to serious secondary infections. Here we demonstrate that measles virus induces tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) mRNA and protein expression in human monocyte-derived dendritic cells. Moreover, measles virus-infected dendritic cells are shown to be cytotoxic via the TRAIL pathway.