bluuebunny/crossref_metadata_2025 huggingface dataset:
Understanding Royal Navy Gas Turbine Sea Water Lubricating Oil Cooler Failures When Caused by Microbial Induced Corrosion (“SRB”)
Authors: Bolwell Richard
Published: November 2005

A managed program to review engine failures and take necessary preventative measures has been in place successfully in the Royal Navy since the introduction of gas turbines into service in the 1970s. One of the more prominent failure mechanisms with the Tyne RM1C and Spey SM1A engines has been the degradation of main line bearings accounting for 25% of all engines rejected. Historically, since the first recorded incident in March 1987, the failures pointed to poor performance of the bearings themselves. However, maintenance studies and recent analysis indicates that a vast proportion have occurred through previously unidentified chloride corrosion as a result of contamination of the lubricating oil system with salt water from the seawater lubricating oil cooler (SWLO cooler). Despite joint ownership of both engine variants with the Royal Netherlands Navy, there was no clear evidence until about five years ago to suggest why tube perforation was occurring. Indeed, the fact that failures have only occurred in Royal Navy service is an interesting twist to the problem. This paper summarizes the phenomenon of SWLO cooler corrosion caused by Microbial Induced Corrosion (principally Sulphate Reducing Bacteria—SRB). It highlights the conditions in which SRB occurs along with demonstrated prevention in Royal Navy gas turbine service through the combined efforts of maintenance and development of a new titanium tubestack. The fault finding and remedial recovery experience may well be of interest to operators of marine gas turbines, both naval and commercial, who use tube type heat exchangers, especially when operating or undertaking work in estuarial waters and nontidal basins or when undertaking littoral duties. This is a practical view of the issue from an operators perspective and while utilizing a wealth of research and technical data available on the subject, it relates the issues at hand to the particular corrosion problem and is not intended as an introduction into organic chemistry.
Inspirations from Biological Optics for Advanced Photonic Systems
Authors: Lee Luke P. , Szema Robert
Published: November 2005

Observing systems in nature has inspired humans to create technological tools that allow us to better understand and imitate biology. Biomimetics, in particular, owes much of its current development to advances in materials science and creative optical system designs. New investigational tools, such as those for microscopic imaging and chemical analyses, have added to our understanding of biological optics. Biologically inspired optical science has become the emerging topic among researchers and scientists. This is in part due to the availability of polymers with customizable optical properties and the ability to rapidly fabricate complex designs using soft lithography and three-dimensional microscale processing techniques.
Retention of Xenon in Quartz and Earth's Missing Xenon
Authors: Sanloup Chrystèle , Schmidt Burkhard C. , Perez Eva Maria Chamorro , Jambon Albert , Gregoryanz Eugene , Mezouar Mohamed
Published: November 2005

The reactivity of xenon with terrestrial oxides was investigated by in situ synchrotron x-ray diffraction. At high temperature (
T
> 500 kelvin), some silicon was reduced, and the pressure stability of quartz was expanded, attesting to the substitution of some xenon for silicon. When the quartz was quenched, xenon diffused out and only a few weight percent remained trapped in samples. These results show that xenon can be covalently bonded to oxygen in quartz in the lower continental crust, providing an answer to the missing xenon problem; synthesis paths of rare gas compounds are also opened.
Bright Infrared Emission from Electrically Induced Excitons in Carbon Nanotubes
Authors: Chen Jia , Perebeinos Vasili , Freitag Marcus , Tsang James , Fu Qiang , Liu Jie , Avouris Phaedon
Published: November 2005

We used the high local electric fields at the junction between the suspended and supported parts of a single carbon nanotube molecule to produce unusually bright infrared emission under unipolar operation. Carriers were accelerated by band-bending at the suspension interface, and they created excitons that radiatively recombined. This excitation mechanism is ∼1000 times more efficient than recombination of independently injected electrons and holes, and it results from weak electron-phonon scattering and strong electron-hole binding caused by one-dimensional confinement. The ensuing high excitation density allows us to observe emission from higher excited states not seen by photoexcitation. The excitation mechanism of these states was analyzed.
Altered TCR Signaling from Geometrically Repatterned Immunological Synapses
Authors: Mossman Kaspar D. , Campi Gabriele , Groves Jay T. , Dustin Michael L.
Published: November 2005

The immunological synapse is a specialized cell-cell junction that is defined by large-scale spatial patterns of receptors and signaling molecules yet remains largely enigmatic in terms of formation and function. We used supported bilayer membranes and nanometer-scale structures fabricated onto the underlying substrate to impose geometric constraints on immunological synapse formation. Analysis of the resulting alternatively patterned synapses revealed a causal relation between the radial position of T cell receptors (TCRs) and signaling activity, with prolonged signaling from TCR microclusters that had been mechanically trapped in the peripheral regions of the synapse. These results are consistent with a model of the synapse in which spatial translocation of TCRs represents a direct mechanism of signal regulation.
Pre- and Postinvasion Defenses Both Contribute to Nonhost Resistance in
Arabidopsis
Authors: Lipka Volker , Dittgen Jan , Bednarek Pawel , Bhat Riyaz , Wiermer Marcel , Stein Monica , Landtag Jörn , Brandt Wolfgang , Rosahl Sabine , Scheel Dierk , Llorente Francisco , Molina Antonio , Parker Jane , Somerville Shauna , Schulze-Lefert Paul
Published: November 2005

Nonhost resistance describes the immunity of an entire plant species against nonadapted pathogen species. We report that
Arabidopsis
PEN2 restricts pathogen entry of two ascomycete powdery mildew fungi that in nature colonize grass and pea species. The PEN2 glycosyl hydrolase localizes to peroxisomes and acts as a component of an inducible preinvasion resistance mechanism. Postinvasion fungal growth is blocked by a separate resistance layer requiring the EDS1-PAD4-SAG101 signaling complex, which is known to function in basal and resistance (
R
) gene–triggered immunity. Concurrent impairment of pre- and postinvasion resistance renders
Arabidopsis
a host for both nonadapted fungi.
Evidence for Macromolecular Protein Rings in the Absence of Bulk Water
Authors: Ruotolo Brandon T. , Giles Kevin , Campuzano Iain , Sandercock Alan M. , Bateman Robert H. , Robinson Carol V.
Published: November 2005

We have examined the architecture of a protein complex in the absence of bulk water. By determining collision cross sections of assemblies of the
trp
RNA binding protein, TRAP, we established that the 11-membered ring topology of the complex can be maintained within a mass spectrometer. We also found that the binding of tryptophan enhances the stability of the ring structure and that addition of a specific RNA molecule increases the size of the complex and prevents structural collapse. These results provide definitive evidence that protein quaternary structure can be maintained in the absence of bulk water and highlight the potential of ion mobility separation for defining shapes of heterogeneous macromolecular assemblies.
Genomic Meanings
Authors: Goldman Michael A.
Published: November 2005

Molecular Models of Life
. Philosophical Papers on Molecular Biology.
By Sahotra Sarkar
. MIT Press, Cambridge, MA, 2005. 412 pp. $38, £24.95. ISBN 0-262-19512-7.

A collection of essays intended to broaden the scope of the philosophy of biology through considerations of molecular biology as a unifying perspective on life.
Porous, Crystalline, Covalent Organic Frameworks
Authors: Côté Adrien P. , Benin Annabelle I. , Ockwig Nathan W. , O'Keeffe Michael , Matzger Adam J. , Yaghi Omar M.
Published: November 2005

Covalent organic frameworks (COFs) have been designed and successfully synthesized by condensation reactions of phenyl diboronic acid {C
6
H
4
[B(OH)
2
]
2
} and hexahydroxytriphenylene [C
18
H
6
(OH)
6
]. Powder x-ray diffraction studies of the highly crystalline products (C
3
H
2
BO)
6
·(C
9
H
12
)
1
(COF-1) and C
9
H
4
BO
2
(COF-5) revealed expanded porous graphitic layers that are either staggered (COF-1,
P
6
3
/
mmc
) or eclipsed (COF-5,
P
6/
mmm
). Their crystal structures are entirely held by strong bonds between B, C, and O atoms to form rigid porous architectures with pore sizes ranging from 7 to 27 angstroms. COF-1 and COF-5 exhibit high thermal stability (to temperatures up to 500° to 600°C), permanent porosity, and high surface areas (711 and 1590 square meters per gram, respectively).
Population Pharmacokinetic Analysis of Vancomycin in Patients with Hematological Malignancies
Authors: Buelga Dolores Santos , del Mar Fernandez de Gatta María , Herrera Emma V. , Dominguez-Gil Alfonso , García María José
Published: November 2005

This study determines vancomycin (VAN) population pharmacokinetics (PK) in adult patients with hematological malignancies. VAN serum concentration data (n= 1,004) from therapeutic drug monitoring were collected retrospectively from 215 patients. A one-compartment PK model was selected. VAN pharmacokinetics population parameters were generated using the NONMEM program. A graphic approach and stepwise generalized additive modeling were used to elucidate the preliminary relationships between PK parameters and clinical covariates analyzed. Covariate selection revealed that total body weight (TBW) affectedV, whereas renal function, estimated by creatinine clearance, and a diagnosis of acute myeloblastic leukemia (AML) influenced VAN clearance. We propose one general and two AML-specific models. The former was defined by CL (liters/h) = 1.08 × CLCR(Cockcroft and Gault)(liters/h); CVCL= 28.16% andV(liters) = 0.98 × TBW; CVV=37.15%. AML models confirmed this structure but with a higher clearance coefficient (1.17). The a priori performance of the models was evaluated in another 59 patients, and clinical suitability was confirmed. The models were fairly accurate, with more than 33% of the measured concentrations being within ±20% of the predicted value. This therapeutic precision is twofold higher than that of a noncustomized population model (16.1%). The corresponding standardized prediction errors included zero and a standard deviation close to unity. The models could be used to estimate appropriate VAN dosage guidelines, which are not clearly defined for this high-risk population. Their simple structure should allow easy implementation in clinical software and application in dosage individualization using the Bayesian approach.
High-Throughput Assays Using a Luciferase-Expressing Replicon, Virus-Like Particles, and Full-Length Virus for West Nile Virus Drug Discovery
Authors: Puig-Basagoiti Francesc , Deas Tia S. , Ren Ping , Tilgner Mark , Ferguson David M. , Shi Pei-Yong
Published: November 2005

Many flaviviruses cause significant human disease worldwide. The development of flavivirus chemotherapy requires reliable high-throughput screening (HTS) assays. Although genetic systems have been developed for many flaviviruses, their usage in antiviral HTS assays has not been well explored. Here we compare three cell-based HTS assays for West Nile virus (WNV) drug discovery: (i) an assay that uses a cell line harboring a persistently replicating subgenomic replicon (containing a deletion of viral structural genes), (ii) an assay that uses packaged virus-like particles containing replicon RNA, and (iii) an assay that uses a full-length reporting virus. A
Renilla
luciferase gene was engineered into the replicon or into the full-length viral genome to monitor viral replication. Potential inhibitors could be identified through suppression of luciferase signals upon compound incubation. The antiviral assays were optimized in a 96-well format, validated with known WNV inhibitors, and proved useful in identifying a new inhibitor(s) through HTS of a compound library. In addition, because each assay encompasses multiple but discrete steps of the viral life cycle, the three systems could potentially be used to discriminate the mode of action of any inhibitor among viral entry (detected by assays ii and iii but not by assay i), replication (including viral translation and RNA synthesis; detected by assays i to iii), and virion assembly (detected by assay iii but not by assays i and ii). The approaches described in this study should be applicable to the development of cell-based assays for other flaviviruses.
Enterococcus faecium
Low-Affinity
pbp5
Is a Transferable Determinant
Authors: Rice Louis B. , Carias Lenore L. , Rudin Susan , Laktičová Viera , Wood Aaron , Hutton-Thomas Rebecca
Published: November 2005

Using 15 unrelated
Enterococcus faecium
isolates as donors, we demonstrated that ampicillin resistance was transferable to an
E. faecium
recipient containing a
pbp5
deletion for all but four strains. The transfers occurred at low frequencies (generally ca. 10
−9
transconjugants/recipient CFU), consistent with chromosome-to-chromosome transfer.
pbp5
transfer occurred within large genetic regions, and insertion into the recipient genome occurred most commonly into the recipient SmaI restriction fragment that had been created by the previous
pbp5
deletion. Restriction mapping of the region upstream of
pbp5
revealed a commonality of fragment sizes among the clinical isolates from the United States which differed significantly from those of three strains that were isolated from turkey feces. These data prove conclusively that
E. faecium pbp5
is a transferable determinant, even in the absence of a coresiding vancomycin resistance mobile element. They also suggest that the spread of high-level ampicillin resistance among U.S.
E. faecium
strains is due in part to the transfer of low-affinity
pbp5
between clinical isolates.
Pharmacodynamics of Caspofungin in a Murine Model of Systemic Candidiasis: Importance of Persistence of Caspofungin in Tissues to Understanding Drug Activity
Authors: Louie Arnold , Deziel Mark , Liu Weiguo , Drusano Michael F. , Gumbo Tawanda , Drusano George L.
Published: November 2005

Pharmacokinetic and pharmacodynamic studies were conducted in a murine model of systemic candidiasis to determine the pharmacodynamic parameter linked with caspofungin efficacy. Additional studies defined the importance of persistent tissue drug concentrations to treatment outcome. The pharmacokinetics of caspofungin were determined in the serum and kidneys of infected mice over 96 h. Population pharmacokinetic analysis demonstrated a serum terminal half-life (t1/2) for caspofungin of 20.2 h when only serum concentrations were considered, but the terminalt1/2increased to 59.2 h when serum and kidney concentration-time data were comodeled. In dose-range studies, the dose-response effect was well described by an inhibitory sigmoid curve for the exposure-effect killing caused by the drug (r2> 0.96;P≪ 0.001). In dose-fractionation studies, fungal counts in kidneys were not statistically different for total doses given as one, two, or four equally divided doses over 96 h, indicating that the area under the concentration-time curve/MIC is the pharmacodynamic parameter that predicts caspofungin efficacy in our infection model. In a separate study, mice infected withCandida albicans24 h after serum concentrations of caspofungin fell below the MIC for the fungal isolate had significant reductions in fungal densities in their kidneys compared with the growth of fungi in the kidneys of untreated controls (P= 0.005). This in vivo biological assay demonstrates that therapeutic concentrations of caspofungin persist at the site of infection in kidney tissue well after serum concentrations fall below the MIC, underscoring the primacy of caspofungin levels in tissues on determining treatment outcome.
Population Pharmacokinetics of Amphotericin B Lipid Complex in Neonates
Authors: Würthwein Gudrun , Groll Andreas H. , Hempel Georg , Adler-Shohet Felice C. , Lieberman Jay M. , Walsh Thomas J.
Published: November 2005

The pharmacokinetics of amphotericin B lipid complex (ABLC) were investigated in neonates with invasive candidiasis enrolled in a phase II multicenter trial. Sparse blood (153 samples; 1 to 9 per patient, 1 to 254 h after the dose) and random urine and cerebrospinal fluid (CSF) samples of 28 neonates (median weight [WT], 1.06 kg; range, 0.48 to 4.9 kg; median gestational age, 27 weeks; range, 24 to 41 weeks) were analyzed. Patients received intravenous ABLC at 2.5 (n= 15) or 5 (n= 13) mg/kg of body weight once a day over 1 or 2 h, respectively, for a median of 21 days (range, 4 to 47 days). Concentrations of amphotericin B were quantified as total drug by high-performance liquid chromatography. Blood data for time after dose (TAD) of <24 h fitted best to a one-compartment model with an additive-error model for residual variability, WT0.75(where 0.75 is an exponent) as a covariate of clearance (CL), and WT as a covariate of volume of distribution (V). Prior amphotericin B, postnatal age, and gestational age did not further improve the model. The final model equations were CL (liters/h) = 0.399 × WT0.75(interindividual variability, 35%) andV(liters) = 10.5 × WT (interindividual variability, 43%). Noncompartmental analysis of pooled data with a TAD of >24 h revealed a terminal half-life of 395 h. Mean concentrations in the urine after 1, 2, and 3 weeks ranged from 0.082 to 0.430 μg/ml, and those in CSF ranged from undetectable to 0.074 μg/ml. The disposition of ABLC in neonates was similar to that observed in other age groups: weight was the only factor that influenced clearance. Based on these results and previously published safety and efficacy data, we recommend a daily dosage between 2.5 and 5.0 mg/kg for treatment of invasiveCandidainfections in neonates.
A Cysteine Protease Inhibitor Protects Dogs from Cardiac Damage during Infection byTrypanosoma cruzi
Authors: Barr S. C. , Warner K. L. , Kornreic B. G. , Piscitelli J. , Wolfe A. , Benet L. , McKerrow J. H.
Published: November 2005

Cruzain is an essential cysteine protease ofTrypanosoma cruziand a therapeutic target for Chagas' disease. Eight dogs were infected withT. cruzi; three were treated with an inhibitor of cruzain, K777, for 14 days. Treatment with K777 abrogated myocardial damage byT. cruzi, as documented by histopathological lesion scores and serum troponin I levels.
Host Restriction ofSalmonella entericaSerotype Typhi Is Not Caused by Functional Alteration of SipA, SopB, or SopD
Authors: Raffatellu Manuela , Sun Yao-Hui , Wilson R. Paul , Tran Quynh T. , Chessa Daniela , Andrews-Polymenis Helene L. , Lawhon Sara D. , Figueiredo Josely F. , Tsolis Renée M. , Adams L. Garry , Bäumler Andreas J.
Published: November 2005

Salmonella entericaserotype Typhi is a strictly human adapted pathogen that does not cause disease in nonprimate vertebrate hosts, whileSalmonella entericaserotype Typhimurium is a broad-host-range pathogen. Serotype Typhi lacks some of the proteins (effectors) exported by the invasion-associated type III secretion system that are required by serotype Typhimurium for eliciting fluid secretion and inflammation in bovine ligated ileal loops. We investigated whether the remaining serotype Typhi effectors implicated in enteropathogenicity (SipA, SopB, and SopD) are functionally exchangeable with their serotype Typhimurium homologues. Serotype Typhi elicited fluid accumulation in bovine ligated ileal loops at levels similar to those elicited by a noninvasive serotype Typhimurium strain (thesipA sopABDE2mutant) or by sterile culture medium. However, introduction of the cloned serotype TyphisipA,sopB, andsopDgenes complemented the ability of a serotype TyphimuriumsipA sopABDE2mutant to elicit fluid secretion in bovine ligated ileal loops. Introduction of the cloned serotype TyphisipA,sopB, andsopDgenes increased the invasiveness of a serotype TyphimuriumsipA sopABDE2mutant for human colon carcinoma epithelial (HT-29 and T84) cells and bovine kidney (MDBK) cells. Translational fusions between the mature TEM-1 β-lactamase reporter and SipA or SopD demonstrated that serotype Typhi translocates these effectors into host cells. We conclude that the inability of serotype Typhi to cause fluid accumulation in bovine ligated ileal loops is not caused by a functional alteration of its SipA, SopB, and SopD effector proteins with respect to their serotype Typhimurium homologues.
β-Arrestin 1 Participates in Platelet-Activating Factor Receptor-Mediated Endocytosis of
Streptococcus pneumoniae
Authors: Radin Jana N. , Orihuela Carlos J. , Murti Gopal , Guglielmo Christopher , Murray Peter J. , Tuomanen Elaine I.
Published: November 2005

Pneumococci traverse eukaryotic cells within vacuoles without intracytoplasmic multiplication. The platelet-activating factor receptor (PAFr) has been suggested as a portal of entry. Pneumococci colocalized with PAFr on endothelial cells and PAFr
−/−
mice showed a substantially impaired ability to support bacterial translocation, particularly from blood to brain. Pneumococci-induced colocalization of PAFr and β-arrestin 1 at the plasma membrane of endothelial cells and PAFr-mediated pneumococcal uptake in transfected COS cells were greatly increased by cotransfection with the scaffold/adapter protein β-arrestin 1. Activation of extracellular signal-regulated kinase kinases was required for uptake and was limited to the cytoplasmic compartment, consistent with activation by β-arrestin rather than PAFr. Uptake of the pneumococcal vacuole involved clathrin, and half the bacteria proceeded into vacuoles marked by Rab5 and later Rab7, the classical route to the lysosome. Overexpression of β-arrestin in endothelial cells decreased colocalization with Rab7. We conclude that the association of β-arrestin with the PAFr contributes to successful translocation of pneumococci.
Helicobacter pylori
Activates Myosin Light-Chain Kinase To Disrupt Claudin-4 and Claudin-5 and Increase Epithelial Permeability
Authors: Fedwick Jason P. , Lapointe Tamia K. , Meddings Jonathan B. , Sherman Philip M. , Buret Andre G.
Published: November 2005

Helicobacter pylori
is a spiral, gram-negative bacterium that specifically and persistently infects the human stomach. In some individuals,
H. pylori
-induced chronic gastritis may progress to gastroduodenal ulcers and gastric cancer. Currently, the host-microbe interactions that determine the clinical outcome of infection are not well defined.
H. pylori
strains capable of disrupting the gastric epithelial barrier may increase the likelihood of developing serious disease. In this study,
H. pylori
strain SS1 increased gastric, but not small intestinal, permeability in C57BL/6 mice.
H. pylori
strain SS1 was able to directly increase paracellular permeability, in the absence of host inflammatory cells, by disrupting the tight-junctional proteins occludin, claudin-4, and claudin-5 in confluent nontransformed epithelial cells.
H. pylori
SS1 also reduced claudin-4 protein levels in human gastric AGS cells. The ability of
H. pylori
SS1 to increase permeability appeared to be independent of the well-characterized virulence factors vacuolating cytotoxin and CagA protein.
H. pylori
activated myosin light-chain kinase in epithelial cells to phosphorylate myosin light chain and increase permeability by disrupting claudin-4 and claudin-5. The bacterial factor responsible for increasing epithelial permeability was heat sensitive, membrane bound, and required apical contact with monolayers. In conclusion, disruptions of the tight junctions observed in this study implicate host cell signaling pathways, including the phosphorylation of myosin light chain and the regulation of tight-junctional proteins claudin-4 and claudin-5, in the pathogenesis of
H. pylori
infection.
Structure, Cellular Distribution, Antigenicity, and Biological Functions of
Fonsecaea pedrosoi
Ceramide Monohexosides
Authors: Nimrichter Leonardo , Cerqueira Mariana D. , Leitão Eduardo A. , Miranda Kildare , Nakayasu Ernesto S. , Almeida Sandro R. , Almeida Igor C. , Alviano Celuta S. , Barreto-Bergter Eliana , Rodrigues Marcio L.
Published: November 2005

Monohexosylceramides (CMHs, or cerebrosides) have been reported as membrane and cell wall constituents of both pathogenic and nonpathogenic fungi, presenting remarkable differences in their ceramide moiety compared to mammalian CMHs. Current evidence suggests that CMHs are involved in fungal differentiation and growth and contribute to host immune response. Here we describe a structural diversity between cerebrosides obtained from different forms of the human pathogen
Fonsecaea pedrosoi
. The major CMH species produced by conidial forms displayed the same structure previously demonstrated by our group for mycelia, an
N
-2′-hydroxyhexadecanoyl-1-β-
d
-glucopyranosyl-9-methyl-4,8-sphingadienine. However, the major cerebroside species purified from sclerotic cells carries an additional hydroxyl group, bound to its long-chain base. The structural difference between cerebrosides from mycelial and sclerotic cells was apparently not relevant for their antigenicity, since they were both recognized at similar levels by sera from individuals with chromoblastomycosis and a monoclonal antibody to a conserved cerebroside structure. Preincubation of fungal cells with anti-CMH monoclonal antibodies had no effect on the interaction of
F. pedrosoi
sclerotic cells with murine macrophages. In contrast to what has been described for other fungal species, sclerotic bodies are resistant to the antifungal action of anti-CMH antibodies. Immunofluorescence analysis showed that recognition of sclerotic cells by these antibodies only occurs at cell wall regions in which melanization is not evident. Accordingly, melanin removal with alkali results in an increased reaction of fungal cells with anti-CMH antibodies. Our results indicate that cerebroside expression in
F. pedrosoi
cells is associated with dimorphism and melanin assembly on the fungal cell wall.
Demonstration of OspC Type Diversity in Invasive Human Lyme Disease Isolates and Identification of Previously Uncharacterized Epitopes That Define the Specificity of the OspC Murine Antibody Response
Authors: Earnhart Christopher G. , Buckles Eric L. , Dumler John Stephen , Marconi Richard T.
Published: November 2005

Outer surface protein C (OspC) of the Lyme disease spirochetes is an important virulence factor that has potential utility for vaccine development. Of the 21 OspC types that have been identified, it has been postulated that types A, B, I, and K are specifically associated with invasive infections. Through an analysis of isolates collected from patients in Maryland we found that OspC types C, D, and N are also associated with invasive infections. This observation suggests that there is greater diversity in the group of OspC types associated with invasive infection than has been previously suggested. Detailed knowledge of the antigenic structure of OspC is essential for vaccine development. To determine if the antibody response to OspC is type specific, recombinant proteins of several different OspC types were immunoblotted and screened with sera from mice infected with isolates having known OspC types. These analyses revealed a high degree of specificity in the antibody response and suggested that the immunodominant epitopes of OspC reside in the variable domains of the protein. To localize these epitopes, OspC fragments were generated and screened with serum collected from infected mice. These analyses led to identification of previously uncharacterized epitopes that define the type specificity of the OspC antibody response. These analyses provide important insight into the antigenic structure of OspC and also provide a basis for understanding the variable nature of the antibody response to this important virulence factor of the Lyme disease spirochetes.
Carbohydrate Moieties of Microsporidian Polar Tube Proteins Are Targeted by Immunoglobulin G in Immunocompetent Individuals
Authors: Peek Ron , Delbac Frédéric , Speijer Dave , Polonais Valérie , Greve Sophie , Wentink-Bonnema Ellen , Ringrose Jeffrey , van Gool Tom
Published: November 2005

Microsporidia of the
Encephalitozoon
species are frequently found as opportunistic pathogens of immunocompromised patients, but very little is known about the prevalence and significance of
Encephalitozoon
infection in immunocompetent individuals. It was reported previously that 8% of Dutch blood donors and 5% of pregnant French women had an immunoglobulin G (IgG) immune response against specific organelles of
Encephalitozoon intestinalis.
These organelles, the so-called polar tube and anchoring disk, are used to penetrate membranes of host cells during infection. The unexpectedly high percentage of immunocompetent individuals with IgG against these organelles suggested that infection of humans with microsporidia might be more common than previously recognized. In the present study, we analyzed this anti-
Encephalitozoon
IgG response by using indirect immunofluorescence, Western blotting, two-dimensional gel electrophoresis, and chemical deglycosylation. Our results show that the antibody response is directed against the posttranslational carbohydrate modification of the major polar tube protein (polar tube protein 1) and carbohydrate moieties of proteins in the anchoring region of the polar tube of
Encephalitozoon
. In addition, the antibodies were found to decrease the infectivity of
E. intestinalis
in vitro. The significance and possible origin of these prevalent antibodies are discussed.
Influence of Pilin Glycosylation on
Pseudomonas aeruginosa
1244 Pilus Function
Authors: Smedley James G. , Jewell Erica , Roguskie Jennifer , Horzempa Joseph , Syboldt Andrew , Stolz Donna Beer , Castric Peter
Published: November 2005

The opportunistic pathogen
Pseudomonas aeruginosa
is a leading cause of nosocomial pneumonia. Among its virulence factors, the type IV pili of
P. aeruginosa
strain 1244 contain a covalently linked, three-sugar glycan of previously unknown significance. The work described in this paper was carried out to determine the influence of the
P. aeruginosa
1244 pilin glycan on pilus function, as well as a possible role in pathogenesis. To accomplish this, a deletion was introduced into the
pilO
gene of this organism. The isogenic knockout strain produced, 1244G7, was unable to glycosylate pilin but could produce pili normal in appearance and quantity. In addition, this strain had somewhat reduced twitching motility, was sensitive to pilus-specific bacteriophages, and could form a normal biofilm. Analysis of whole cells and isolated pili from wild-type
P. aeruginosa
strain 1244 by transmission electron microscopy with a glycan-specific immunogold label showed that this saccharide was distributed evenly over the fiber surface. The presence of the pilin glycan reduced the hydrophobicity of purified pili as well as whole cells. With regard to pathogenicity,
P. aeruginosa
strains producing glycosylated pili were commonly found among clinical isolates and particularly among those strains isolated from sputum. Competition index analysis using a mouse respiratory model comparing strains 1244 and 1244G7 indicated that the presence of the pilin glycan allowed for significantly greater survival in the lung environment. These results collectively suggest that the pilin glycan is a significant virulence factor and may aid in the establishment of infection.
Capsule-Negative
Staphylococcus aureus
Induces Chronic Experimental Mastitis in Mice
Authors: Tuchscherr Lorena P. N. , Buzzola Fernanda R. , Alvarez Lucía P. , Caccuri Roberto L. , Lee Jean C. , Sordelli Daniel O.
Published: November 2005

Staphylococcus aureus
capsular polysaccharides (CP) have been shown to enhance staphylococcal virulence in numerous animal models of infection. Although serotype 5 CP (CP5) and CP8 predominate among
S. aureus
isolates from humans, most staphylococcal isolates from bovines with mastitis in Argentina are capsule negative. This study was designed to evaluate the effects of CP5 and CP8 expression on the pathogenesis of experimental murine mastitis. Lactating mice were challenged by the intramammary route with one of three isogenic
S. aureus
strains producing CP5, CP8, or no capsule. Significantly greater numbers of acapsular mutant cells were recovered from the infected glands 12 days after bacterial challenge compared with the encapsulated strains. Histopathological analyses revealed greater polymorphonuclear and mononuclear leukocyte infiltration and congestion in the mammary glands of mice infected with the encapsulated strains compared with the acapsular mutant, and the serotype 5 strain elicited more inflammation than the serotype 8 strain. In vitro experiments revealed that the acapsular
S. aureus
strain was internalized by MAC-T bovine epithelial cells in significantly greater numbers than the CP5- or CP8-producing strain. Taken together, the results suggest that
S. aureus
lacking a capsule was able to persist in the murine mammary gland, whereas encapsulated strains elicited more inflammation and were eliminated faster. Loss of CP5 or CP8 expression may enhance the persistence of staphylococci in the mammary glands of chronically infected hosts.
A Leucine-Rich Motif Targets
Pseudomonas aeruginosa
ExoS within Mammalian Cells
Authors: Zhang Yue , Barbieri Joseph T.
Published: November 2005

Type III cytotoxins contribute to the ability of bacterial pathogens to subvert the host innate immune system. ExoS (453 amino acids) is a bifunctional type III cytotoxin produced by
Pseudomonas aeruginosa
. Residues 96 to 232 comprise a Rho GTPase activating protein domain, while residues 233 to 453 comprise a 14-3-3-dependent ADP-ribosyltransferase domain. An N-terminal domain (termed the membrane localization domain [MLD]) targets ExoS to the Golgi-endoplasmic reticulum (Golgi-ER) of mammalian cells. This study identifies an amino acid motif that is responsible for the membrane binding properties of the MLD. Deletion mapping showed that the MLD included a symmetrical leucine-rich motif within residues 51 to 77 of ExoS. The terminal dileucines and internal leucines and an isoleucine within the MLD, but not charged or other hydrophobic residues, targeted a reporter protein to the Golgi-ER region of HeLa cells. Mutations of the leucines within the MLD did not affect type III secretion or translocation into HeLa cells but limited the ability of ExoS to ADP-ribosylate Ras GTPases. Mutations of charged residues within the MLD did not affect type III secretion, delivery into HeLa cells, or the ability of ExoS to ADP-ribosylate Ras GTPases. The organization of the leucines within the MLD of ExoS is different from that of previously described leucine-rich motifs but is present in several other bacterial proteins. This implies a role for intracellular targeting in the efficient targeting of mammalian cells by type III cytotoxins.
Neonatal and Maternal Immunological Responses to Conserved Epitopes within the DBL-γ3 Chondroitin Sulfate A-Binding Domain ofPlasmodium falciparumErythrocyte Membrane Protein 1
Authors: Brustoski Kim , Kramer Martin , Möller Ulrike , Kremsner Peter G. , Luty Adrian J. F.
Published: November 2005

Plasmodium falciparumerythrocyte membrane protein 1 (PfEMP1) mediates the adherence ofP. falciparum-infected erythrocytes to placental syncytiotrophoblasts via interactions with chondroitin sulfate A (CSA), a characteristic of pregnancy-associated malaria. Pregnancy-associated malaria predicts increased susceptibility of newborns to malaria, and it is postulated that transplacental passage of parasite antigen induces immune regulatory activity in the neonate. We wished to examine the immune responsiveness to a CSA-binding domain of PfEMP1, the DBL-γ3 domain, in cord and maternal venous blood obtained from pregnancies with various histories ofP. falciparuminfection. We assessed in vitro T-cell cytokine and plasma immunoglobulin G (IgG) and IgM responses to four peptides corresponding to highly conserved regions of a DBL-γ3 domain common to central African parasite isolates. The presence of placentalP. falciparuminfection at delivery was associated with elevated frequencies of DBL-γ3 peptide-specific CD3+interleukin-10-positive T cells in cord blood, while treatment and clearance of infection prior to delivery was associated with elevated frequencies of CD3+gamma interferon-positive T cells. DBL-γ3 peptide-specific IgM antibodies were detected in 12 of 60 (20%) cord plasma samples from those born to mothers withP. falciparuminfection during pregnancy. Consistent with polyclonal anti-PfEMP1 antibody responses that are associated with protection against pregnancy-associated malaria, the presence of maternal IgG antibodies with specificity for one of the DBL-γ3 peptides showed a parity-dependent profile. These data demonstrate that peptides corresponding to conserved regions of the DBL-γ3 domain of PfEMP1 are immunogenic inP. falciparum-infected mothers and their offspring.
Phase I Malaria Vaccine Trial with a Long Synthetic Peptide Derived from the Merozoite Surface Protein 3 Antigen
Authors: Audran Régine , Cachat Michel , Lurati Floriana , Soe Soe , Leroy Odile , Corradin Giampietro , Druilhe Pierre , Spertini François
Published: November 2005

The C-terminal conserved region ofPlasmodium falciparummerozoite surface protein 3 (MSP3) is the trigger antigen of a protective immune response mediated by cytophilic antibodies. In an open, randomized, two-adjuvant (Montanide ISA 720, aluminum hydroxide) phase I clinical trial we evaluated the safety and immunogenicity of increasing doses of a long synthetic peptide construct spanning the conserved region of MSP3 targeted by biologically active antibodies (MSP3-LSP). Thirty-five healthy volunteers were randomized to receive three subcutaneous injections on days 0, 30, and 120. Of the 100 injections given, 10 caused severe local reactions, 62 caused transient mild to moderate local reactions, and 28 caused no reaction. On the basis of preestablished exclusion criteria, use of the Montanide formulation led to withdrawal of five volunteers after the second injection. This led to a reduction in the subsequent vaccine doses in four of the groups. No vaccine-related serious adverse events occurred throughout the trial. After the third injection, volunteers displayed a marked specific anti-MSP3-LSP antibody response (23/30 individuals, compared with 29/34 individuals for plasma from an area where malaria is endemic), an anti-native MSP3 antibody response (19/30 individuals), a T-cell-antigen-specific proliferative response (26/30 individuals), and gamma interferon production (25/30 individuals). In conclusion, the MSP3-LSP vaccine was immunogenic with both adjuvants, although it was unacceptably reactogenic when it was combined with Montanide. The potential usefulness of the candidate vaccine is supported by the induction of a strong cytophilic response (i.e., the type of anti-MSP3 antibodies involved in antibody-dependent, monocyte-mediated protective mechanisms in areas where malaria is endemic).
Community-Based Safety, Immunogenicity, and Transmissibility Study of the
Shigella sonnei
WRSS1 Vaccine in Israeli Volunteers
Authors: Orr Nadav , Katz David E. , Atsmon Jacob , Radu Paull , Yavzori Miri , Halperin Tamar , Sela Tamar , Kayouf Raid , Klein Zivit , Ambar Ruhama , Cohen Dani , Wolf Marcia K. , Venkatesan Malabi M. , Hale Thomas L.
Published: November 2005

We describe the first community-based evaluation of
Shigella sonnei
strain WRSS1, a live, oral candidate vaccine attenuated by a 212-bp deletion in the
virG
(or
icsA
) plasmid virulence gene. Three single-dose regimens of WRSS1 (5 × 10
3
CFU, 2 × 10
4
CFU, and 4 × 10
5
CFU) were tested with cohorts of 15 adult volunteers. The vaccine was generally well tolerated at the 10
3
- and 10
4
-CFU doses. There were no fevers and there was one report of moderate diarrhea in 30 vaccinees; five additional vaccinees reported mild diarrhea. At the 10
5
-CFU dose, there were two reports of low-grade fevers and four reports of moderate diarrhea. The geometric means for immunoglobulin A (IgA) antibody-secreting cells (ASC) against lipopolysaccharide (LPS) were 30, 75, and 193 ASC per 10
6
peripheral blood mononuclear cells (PBMC) for the 10
3
-, 10
4
-, and 10
5
-CFU doses, respectively. The IgG means were 40, 46, and 135 ASC per 10
6
PBMC, respectively. The 10
4
-CFU dose of WRSS1 gave the best balance of safety and immunogenicity, since all vaccinees had a significant IgA ASC response and 73% had a response of more than 50 ASC. The anti-LPS seroconversion rate (threefold) for IgA was 60% and the IgG rate was 27% for the 10
4
-CFU cohort. Each vaccinee and a cohabitating household contact delivered daily perianal stool swabs for bacteriological culture. WRSS1 colonized vaccinees for a median of 5 days, and one individual excreted WRSS1 intermittently for 23 days. None of the 45 household contacts were colonized with WRSS1 after a cumulative 192 days of cohabitation with colonized vaccinees, suggesting that adventitious vaccine spread was not common in the community setting.
DltABCD- and MprF-Mediated Cell Envelope Modifications of
Staphylococcus aureus
Confer Resistance to Platelet Microbicidal Proteins and Contribute to Virulence in a Rabbit Endocarditis Model
Authors: Weidenmaier Christopher , Peschel Andreas , Kempf Volkhard A. J. , Lucindo Natalie , Yeaman Michael R. , Bayer Arnold S.
Published: November 2005

The DltABCD and MprF proteins contribute a net positive charge to the
Staphylococcus aureus
surface envelope by alanylating and lysinylating teichoic acids and membrane phosphatidylglycerol, respectively. These surface charge modifications are associated with increased in vitro resistance profiles of
S. aureus
to a number of endogenous cationic antimicrobial peptides (CAPs), such as α-defensins. The current study investigated the effects of
dltA
and
mprF
mutations on the following host factors relevant to endovascular infections: (i) in vitro susceptibility to the CAP thrombin-induced platelet microbicidal protein 1 (tPMP-1), (ii) in vitro adherence to endothelial cells (EC) and matrix proteins, and (iii) in vivo virulence in an endovascular infection model (rabbit endocarditis) in which tPMP-1 is felt to play a role in limiting
S. aureus
pathogenesis. Both mutations resulted in substantial increases in the in vitro susceptibility to tPMP-1 compared to that of the parental strain. The
dltA
(but not the
mprF
) mutation resulted in a significantly reduced capacity to bind to EC in vitro, while neither mutation adversely impacted in vitro binding to fibronectin, fibrinogen, or platelets. In vivo, both mutations significantly attenuated virulence in terms of early colonization of sterile vegetations and subsequent proliferation at this site (versus the parental strain). However, only the
dltA
mutation significantly reduced metastatic infections in kidneys and spleens compared to those in animals infected with the parental strain. These data underscore the importance of resistance to distinct CAPs and of teichoic acid-dependent EC interactions in the context of endovascular infection pathogenesis.
Mouse Intestine Selects Nonmotile
flhDC
Mutants of
Escherichia coli
MG1655 with Increased Colonizing Ability and Better Utilization of Carbon Sources
Authors: Leatham Mary P. , Stevenson Sarah J. , Gauger Eric J. , Krogfelt Karen A. , Lins Jeremy J. , Haddock Traci L. , Autieri Steven M. , Conway Tyrrell , Cohen Paul S.
Published: November 2005

d
-Gluconate which is primarily catabolized via the Entner-Doudoroff (ED) pathway, has been implicated as being important for colonization of the streptomycin-treated mouse large intestine by
Escherichia coli
MG1655, a human commensal strain. In the present study, we report that an MG1655 Δ
edd
mutant defective in the ED pathway grows poorly not only on gluconate as a sole carbon source but on a number of other sugars previously implicated as being important for colonization, including
l
-fucose,
d
-gluconate,
d
-glucuronate,
N
-acetyl-
d
-glucosamine,
d
-mannose, and
d
-ribose. Furthermore, we show that the mouse intestine selects mutants of MG1655 Δ
edd
and wild-type MG1655 that have improved mouse intestine-colonizing ability and grow 15 to 30% faster on the aforementioned sugars. The mutants of MG1655 Δ
edd
and wild-type MG1655 selected by the intestine are shown to be nonmotile and to have deletions in the
flhDC
operon, which encodes the master regulator of flagellar biosynthesis. Finally, we show that Δ
flhDC
mutants of wild-type MG1655 and MG1655 Δ
edd
constructed in the laboratory act identically to those selected by the intestine; i.e., they grow better than their respective parents on sugars as sole carbon sources and are better colonizers of the mouse intestine.
Virulence and Karyotype Analyses of
rad52
Mutants of
Candida albicans
: Regeneration of a TruncatedChromosome of a Reintegrant Strain (
rad52
/
RAD52
) in the Host
Authors: Chauhan Neeraj , Ciudad Toni , Rodríguez-Alejandre Ane , Larriba Germán , Calderone Richard , Andaluz Encarnación
Published: November 2005

The
virulence of
Candida albicans
mutants lacking one or both
copies of
RAD52
, a gene involved in homologous recombination
(HR), was evaluated in a murine model of hematogenously disseminated
candidiasis. In this study, the virulence of the
rad52
Δ mutant was dependent upon the inoculum
concentration. Mice survived at a cell inoculum of 1 ×
10
6
, but there was a decrease in survival time at dosages of
1.5 × 10
6
and especially at 3 ×
10
6
cells per animal. The heterozygote
RAD52/rad52
behaved like wild type, whereas a reintegrant strain was intermediate
in its ability to cause death compared to these strains and to the
avirulent
rad52/rad52
null at inocula of 1 ×
10
6
and 1.5 × 10
6
cells. A double mutant,
lig4/lig4/rad52/rad52
, was avirulent at all inocula used. PCR
analysis of the
RAD52
and/or
LIG4
loci showed that
all strains recovered from animals matched the genotype of the
inoculated strains. Analysis of the electrophoretical karyotypes
indicated that the inoculated, reintegrant strain carried a large
deletion in one copy of chromosome 6 (the shortest homologue, or
Chr6b). Interestingly, truncated Chr6b was regenerated in all the
strains recovered from moribund animals using the homologue as a
template. Further, regeneration of Chr6b was paralleled by an increase
in virulence that was still lower than that of wild type, likely
because of the persistent loss of heterozygosity in the regenerated
region. Overall, our results indicate that systemic candidiasis can
develop in the absence of HR, but simultaneous elimination of both
recombination pathways, HR and nonhomologous end-joining, suppresses
virulence even at very high
inocula.