bluuebunny/crossref_metadata_2025 huggingface dataset:
Binding to pyruvylated compounds as an ancestral mechanism to anchor the outer envelope in primitive bacteria
Authors: Cava Felipe , De Pedro Miguel A. , Schwarz Heinz , Henne Anke , Berenguer José
Published: April 2004

Electron microscopy of isolated cell walls of the ancient bacterium Thermus thermophilus revealed that most of the peptidoglycan (PG) surface, apart from the septal region, was shielded against specific αPG antibodies. On the other hand, an antiserum raised against S‐layer‐attached cell wall fragments (αSAC) bound to most of the surface except for the septal regions. Treatments with α‐amylase and pronase E made the entire cell wall surface uniformly accessible to αPG and severely decreased the binding of αSAC. We concluded that a layer of strongly bound secondary cell wall polymers (SCWPs) covers most of the cell wall surface in this ancient bacterium. A preliminary analysis revealed that such SCWPs constitute 14% of the cell wall and are essentially composed of sugars. Enzyme treatments of the cell walls revealed that SCWP was required in vitro for the binding of the S‐layer protein through the S‐layer homology (SLH) motif. The csaB gene was necessary for the attachment of the S‐layer–outer membrane (OM) complex to the cell wall in growing cells of T. thermophilus. In vitro experiments confirmed that cell walls from a csaB mutant bound to the S‐layer with a much lower affinity (∼1/10) than that of the wild type. CsaB was found to be required for pyruvylation of components of the SCWP and for immunodetection with α‐SAC antiserum. Therefore, the S‐layer–OM complex of T. thermophilus binds to the cell wall through the SLH motif of the S‐layer protein via a strong interaction with a highly immunogenic pyruvylated component of the SCWP. Immuno‐cross‐reactive compounds were detected with αSAC on cell walls of other Thermus spp. and in the phylogenetically related microorganism Deinococcus radiodurans. These results imply that the interaction between the SLH motif and pyruvylated components of the cell wall arose early during bacterial evolution as an ancestral mechanism for anchoring proteins and outer membranes to the cell walls of primitive bacteria.
Natural transformation and phase variation modulation in Neisseria meningitidis
Authors: Alexander Heather L. , Richardson Anthony R. , Stojiljkovic Igor
Published: April 2004

Neisseria meningitidis has evolved the ability to control the expression‐state of numerous genes by phase variation. It has been proposed that the process aids this human pathogen in coping with the diversity of microenvironments and host immune systems. Therefore, increased frequencies of phase variation may augment the organism's adaptability and virulence. In this study, we found that DNA derived from various neisserial co‐colonizers of the human nasopharynx increased N. meningitidis switching frequencies, indicating that heterologous neisserial DNA modulates phase variation in a transformation‐dependent manner. In order to determine whether the effect of heterologous DNA was specific to the Hb receptor, HmbR, we constructed a Universal Rates of Switching cassette (UROS). With this cassette, we demonstrated that heterologous DNA positively affects phase variation throughout the meningococcal genome, as UROS phase variation frequencies were also increased in the presence of neisserial DNA. Overexpressing components of the neisserial mismatch repair system partially alleviated DNA‐induced changes in phase variation frequencies, thus implicating mismatch repair titration as a cause of these transformation‐dependent increases in switching. The DNA‐dependent effect on phase variation was transient and may serve as a mechanism for meningococcal genetic variability that avoids the fitness costs encountered by global mutators.
EzrA prevents aberrant cell division by modulating assembly of the cytoskeletal protein FtsZ
Authors: Haeusser Daniel P. , Schwartz Rachel L. , Smith Alison M. , Oates Michelle Erin , Levin Petra Anne
Published: April 2004

In response to a cell cycle signal, the cytoskeletal protein FtsZ assembles into a ring structure that establishes the location of the division site and serves as a framework for assembly of the division machinery. A battery of factors control FtsZ assembly to ensure that the ring forms in the correct position and at the precise time. EzrA, a negative regulator of FtsZ ring formation, is important for ensuring that the ring forms only once per cell cycle and that cytokinesis is restricted to mid‐cell. EzrA is distributed throughout the plasma membrane and localizes to the ring in an FtsZ‐dependent manner, suggesting that it interacts directly with FtsZ to modulate assembly. We have performed a series of experiments examining the interaction between EzrA and FtsZ. As little as twofold overexpression of EzrA blocks FtsZ ring formation in a sensitized genetic background, consistent with its predicted function. A purified EzrA fusion protein interacts directly with FtsZ to block assembly in vitro. Although EzrA is able to inhibit FtsZ assembly, it is unable to disassemble preformed polymers. These data support a model in which EzrA interacts directly with FtsZ at the plasma membrane to prevent polymerization and aberrant FtsZ ring formation.
A suf operon requirement for Fe–S cluster assembly during iron starvation in Escherichia coli
Authors: Outten F. Wayne , Djaman Ouliana , Storz Gisela
Published: April 2004

The suf and isc operons of Escherichia coli have been implicated in Fe–S cluster assembly. However, it has been unclear why E. coli has two systems for Fe–S cluster biosynthesis. We have examined the regulatory characteristics and mutant phenotypes of both operons to discern if the two operons have redundant functions or if their cellular roles are divergent. Both operons are similarly induced by hydrogen peroxide and the iron chelator 2,2′‐dipyridyl, although by different mechanisms. Regulation of the isc operon is mediated by IscR, whereas the suf operon requires OxyR and IHF for the response to oxidative stress and Fur for induction by iron starvation. Simultaneous deletion of iscS and most suf genes is synthetically lethal. However, although the suf and isc operons have overlapping functions, they act as distinct complexes because the SufS desulphurase alone cannot substitute for the IscS enzyme. In addition, suf deletion mutants are more sensitive to iron starvation than isc mutants, and the activity of the Fe–S enzyme gluconate dehydratase is diminished in the suf mutant during iron starvation. These findings are consistent with the model that the isc operon encodes the housekeeping Fe–S cluster assembly system in E. coli, whereas the suf operon is specifically adapted to synthesize Fe–S clusters when iron or sulphur metabolism is disrupted by iron starvation or oxidative stress.
Characterization of a complex chemosensory signal transduction system which controls twitching motility in Pseudomonas aeruginosa
Authors: Whitchurch Cynthia B. , Leech Andrew J. , Young Michael D. , Kennedy Derek , Sargent Jennifer L. , Bertrand Jacob J. , Semmler Annalese B. T. , Mellick Albert S. , Martin Paul R. , Alm Richard A. , Hobbs Matthew , Beatson Scott A. , Huang Bixing , Nguyen Lam , Commolli James C. , Engel Joanne N. , Darzins Aldis , Mattick John S.
Published: April 2004

Virulence of the opportunistic pathogen Pseudomonas aeruginosa involves the coordinate expression of a wide range of virulence factors including type IV pili which are required for colonization of host tissues and are associated with a form of surface translocation termed twitching motility. Twitching motility in P. aeruginosa is controlled by a complex signal transduction pathway which shares many modules in common with chemosensory systems controlling flagella rotation in bacteria and which is composed, in part, of the previously described proteins PilG, PilH, PilI, PilJ and PilK. Here we describe another three components of this pathway: ChpA, ChpB and ChpC, as well as two downstream genes, ChpD and ChpE, which may also be involved. The central component of the pathway, ChpA, possesses nine potential sites of phosphorylation: six histidine‐containing phosphotransfer (HPt) domains, two novel serine‐ and threonine‐containing phosphotransfer (SPt, TPt) domains and a CheY‐like receiver domain at its C‐terminus, and as such represents one of the most complex signalling proteins yet described in nature. We show that the Chp chemosensory system controls twitching motility and type IV pili biogenesis through control of pili assembly and/or retraction as well as expression of the pilin subunit gene pilA. The Chp system is also required for full virulence in a mouse model of acute pneumonia.
The molecular basis of fibronectin‐mediated bacterial adherence to host cells
Authors: Schwarz‐Linek Ulrich , Höök Magnus , Potts Jennifer R.
Published: April 2004

Many pathogenic Gram‐positive bacteria produce cell wall‐anchored proteins that bind to components of the extracellular matrix (ECM) of the host. These bacterial MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) are thought to play a critical role in infection. One group of MSCRAMMs, produced by staphylococci and streptococci, targets fibronectin (Fn, a glycoprotein found in the ECM and body fluids of vertebrates) using repeats in the C‐terminal region of the bacterial protein. These bacterial Fn‐binding proteins (FnBPs) mediate adhesion to host tissue and bacterial uptake into non‐phagocytic host cells. Recent studies on interactions between the host and bacterial proteins at the residue‐specific level and on the mechanism of host cell invasion are providing a much clearer picture of these processes.
Seed‐specific expression of a bacterial phosphoenolpyruvate carboxylase inVicia narbonensisincreases protein content and improves carbon economy
Authors: Rolletschek Hardy , Borisjuk Ljudmilla , Radchuk Ruslana , Miranda Manoela , Heim Ute , Wobus Ulrich , Weber Hans
Published: April 2004

An ambitious aim in plant breeding and biotechnology is to increase the protein content of crop seeds used for food and feed. Using an approach to manipulate assimilate partitioning, we succeeded in elevating the protein content in legume seeds up to 50%. Transgenic bean plants were generated which express aCorynebacterium glutamicumphosphoenolpyruvate carboxylase (PEPC) in a seed‐specific manner. The bacterial enzyme was not feedback inhibited by malate. Transgenic seeds showed a higher [14C]‐CO2uptake and about a threefold increased incorporation of labelled carbon into proteins. Changed metabolite profiles of maturing cotyledons indicated a shift of metabolic fluxes from sugars/starch into organic acids and free amino acids. These changes were consistent with an increased carbon flow through the anaplerotic pathway catalysed by PEPC. Consequently, transgenic seeds accumulated up to 20% more protein per gram seed dry weight. Additionally, seed dry weight was higher by 20%−30%. We conclude that PEPC in seeds is a promising target for molecular plant breeding.
Efficient cloning of plant genomes into bacterial artificial chromosome (BAC) libraries with larger and more uniform insert size
Authors: Chalhoub Boulos , Belcram Harry , Caboche Michel
Published: April 2004

The construction of bacterial artificial chromosome (BAC) libraries remains relatively complex and laborious, such that any technological improvement is considered to be highly advantageous. In this study, we addressed several aspects that improved the quality and efficiency of cloning of plant genomes into BACs. We set the ‘single tube vector’ preparation method with no precipitation or gel electrophoresis steps, which resulted in less vector DNA damage and a remarkable two‐ to threefold higher transformation efficiency compared with other known vector preparation methods. We used a reduced amount of DNA for partial digestion (up to 5 µg), which resulted in less BAC clones with small inserts. We performed electrophoresis in 0.25 × TBE (Tris, boric acid, ethylenediaminetetraacetic acid) buffer instead of 0.5 × TBE, which resulted in larger and more uniformly sized BAC inserts and, surprisingly, a two‐ to threefold higher transformation efficiency, probably due to less contamination with borate ions. We adopted a triple size selection that resulted in an increased mean insert size of up to 70 kb and a transformation efficiency comparable with that of double size selection. Overall, the improved protocol presented in this study resulted in a five‐ to sixfold higher cloning efficiency and larger and more uniformly sized BAC inserts. BAC libraries with the desired mean insert size (up to 200 kb) were constructed from several plant species, including hexaploid wheat. The improved protocol will render the construction of BAC libraries more available in plants and will greatly enhance genome analysis, gene mapping and cloning.
Expression of Vitreoscilla haemoglobin in tobacco cell cultures relieves nitrosative stress in vivo and protects from NO in vitro
Authors: Frey Alexander D. , Oberle Bettina T. , Farrés Judith , Kallio Pauli T.
Published: April 2004

Targeted expression of Vitreoscilla haemoglobin (VHb) has been analysed in Nicotiana tabacum plants and suspension cultures under various growth and stress conditions. VHb localization to different cell compartments (cytoplasm, chloroplast and mitochondria) was successful, as judged by signal peptide cleavage. The presence of VHb in subcellular compartments did not result in phenotypical differences between these plant lines. In contrast with previous reports, we were unable to discern any significant changes in growth and other phenotypical characteristics between VHb‐expressing and transformed control plants under standard growth conditions. When exposed to nitrosative stress, growth of VHb‐expressing cultures was less affected relative to transformed controls. Furthermore, a diminished inactivation of the NO‐sensitive enzyme aconitase was observed in the presence of VHb. In contrast, no protective effect of VHb expression against oxidative stress could be detected.
Inflammation as a cause of malnutrition, atherosclerotic cardiovascular disease, and poor outcome in hemodialysis patients
Authors: Yao Qiang , Lindholm Bengt , Stenvinkel Peter
Published: April 2004

Cardiovascular disease (CVD) remains the major cause of morbidity and mortality in end‐stage renal disease (ESRD) patients treated by hemodialysis (HD). Although traditional risk factors are common in dialysis patients, they may not alone be sufficient to account for the unacceptable high prevalence of CVD in this patient group. Recent evidence demonstrates that chronic inflammation, a nontraditional risk factor that is commonly observed in HD patients, may cause malnutrition and progressive atherosclerotic CVD by several pathogenetic mechanisms. The cause(s) of inflammation in HD patients is multifactorial and includes both dialysis‐related (such as graft and fistula infections, bioincompatibility, impure dialysate, and back‐filtration) and dialysis‐unrelated factors. Although inflammation may reflect underlying CVD, an acute‐phase reaction may also be a direct cause of vascular injury. Available data suggest that proinflammatory cytokines play a central role in the genesis of both malnutrition and CVD in ESRD. Thus, it could be speculated that suppression of the vicious cycle of malnutrition, inflammation, and atherosclerosis (MIA syndrome) would improve survival in dialysis patients. As there is not yet any recognized, or even proposed, targeted treatment for ESRD patients with chronic inflammation; it would be of considerable interest to study the long‐term effect of various anti‐inflammatory treatment strategies on nutritional and cardiovascular status as well as outcome in these patients.
The Physics of Neutron Stars
Authors: Lattimer J. M. , Prakash M.
Published: April 2004

Neutron stars are some of the densest manifestations of massive objects in the universe. They are ideal astrophysical laboratories for testing theories of dense matter physics and provide connections among nuclear physics, particle physics, and astrophysics. Neutron stars may exhibit conditions and phenomena not observed elsewhere, such as hyperon-dominated matter, deconfined quark matter, superfluidity and superconductivity with critical temperatures near 10
10
kelvin, opaqueness to neutrinos, and magnetic fields in excess of 10
13
Gauss. Here, we describe the formation, structure, internal composition, and evolution of neutron stars. Observations that include studies of pulsars in binary systems, thermal emission from isolated neutron stars, glitches from pulsars, and quasi-periodic oscillations from accreting neutron stars provide information about neutron star masses, radii, temperatures, ages, and internal compositions.
Self-Assembly of Ordered, Robust, Three-Dimensional Gold Nanocrystal/Silica Arrays
Authors: Fan Hongyou , Yang Kai , Boye Daniel M. , Sigmon Thomas , Malloy Kevin J. , Xu Huifang , López Gabriel P. , Brinker C. Jeffrey
Published: April 2004

We report the synthesis of a new nanocrystal (NC) mesophase through self-assembly of water-soluble NC micelles with soluble silica. The mesophase comprises gold nanocrystals arranged within a silica matrix in a face-centered cubic lattice with cell dimensions that are adjustable through control of the nanocrystal diameter and/or the alkane chain lengths of the primary alkanethiol stabilizing ligands or the surrounding secondary surfactants. Under kinetically controlled silica polymerization conditions, evaporation drives self-assembly of NC micelles into ordered NC/silica thin-film mesophases during spin coating. The intermediate NC micelles are water soluble and of interest for biolabeling. Initial experiments on a metal-insulator-metal capacitor fabricated with an ordered three-dimensional gold nanocrystal/silica array as the “insulator” demonstrated collective Coulomb blockade behavior below 100 kelvin and established the current-voltage scaling relationship for a well-defined three-dimensional array of Coulomb islands.
The Rise of the Rhizosolenid Diatoms
Authors: Damsté Jaap S. Sinninghe , Muyzer Gerard , Abbas Ben , Rampen Sebastiaan W. , Massé Guillaume , Allard W. Guy , Belt Simon T. , Robert Jean-Michel , Rowland Steven J. , Moldowan J. Michael , Barbanti Silvana M. , Fago Frederick J. , Denisevich Peter , Dahl Jeremy , Trindade Luiz A. F. , Schouten Stefan
Published: April 2004

The 18S ribosomal DNA molecular phylogeny and lipid composition of over 120 marine diatoms showed that the capability to biosynthesize highly branched isoprenoid (HBI) alkenes is restricted to two specific phylogenetic clusters, which independently evolved in centric and pennate diatoms. The molecular record of C
25
HBI chemical fossils in a large suite of well-dated marine sediments and petroleum revealed that the older cluster, composed of rhizosolenid diatoms, evolved 91.5 ± 1.5 million years ago (Upper Turonian), enabling an accurate dating of the pace of diatom evolution that is unprecedented. The rapid rise of the rhizosolenid diatoms probably resulted from a major reorganization of the nutrient budget in the mid-Cretaceous oceans, triggered by plate tectonics.
Biological Effects of Short-Term or Prolonged Administration of 9-[2-(Phosphonomethoxy)Propyl]Adenine (Tenofovir) to Newborn and Infant Rhesus Macaques
Authors: Van Rompay Koen K. A. , Brignolo Laurie L. , Meyer Dennis J. , Jerome Christopher , Tarara Ross , Spinner Abigail , Hamilton Marta , Hirst Linda L. , Bennett David R. , Canfield Don R. , Dearman Trish G. , Von Morgenland Wilhelm , Allen Phil C. , Valverde Celia , Castillo Alesha B. , Martin R. Bruce , Samii Valerie F. , Bendele Ray , Desjardins John , Marthas Marta L. , Pedersen Niels C. , Bischofberger Norbert
Published: April 2004

The reverse transcriptase inhibitor 9-[2-(phosphonomethoxy)propyl]adenine (PMPA; tenofovir) was previously found to offer strong prophylactic and therapeutic benefits in an infant macaque model of pediatric human immunodeficiency virus (HIV) infection. We now summarize the toxicity and safety of PMPA in these studies. When a range of PMPA doses (4 to 30 mg/kg of body weight administered subcutaneously once daily) was administered to 39 infant macaques for a short period of time (range, 1 day to 12 weeks), no adverse effects on their health or growth were observed; this included a subset of 12 animals which were monitored for more than 2 years. In contrast, daily administration of a high dose of PMPA (30 mg/kg subcutaneously) for prolonged periods of time (>8 to 21 months) to 13 animals resulted in a Fanconi-like syndrome (proximal renal tubular disorder) with glucosuria, aminoaciduria, hypophosphatemia, growth restriction, bone pathology (osteomalacia), and reduced clearance of PMPA. The adverse effects were reversible or were alleviated following either complete withdrawal of PMPA treatment or reduction of the daily regimen from 30 mg/kg to 2.5 to 10 mg/kg subcutaneously. Finally, to evaluate the safety of a prolonged low-dose treatment regimen, two newborn macaques were started on a 10-mg/kg/day subcutaneous regimen; these animals are healthy and have normal bone density and growth after 5 years of daily treatment. In conclusion, our findings suggest that chronic daily administration of a high dose of PMPA results in adverse effects on kidney and bone, while short-term administration of relatively high doses and prolonged low-dose administration are safe.
Variety of β-Lactamases Produced by Amoxicillin-Clavulanate-Resistant
Escherichia coli
Isolated in the Northeastern United States
Authors: Kaye Keith S. , Gold Howard S. , Schwaber Mitchell J. , Venkataraman Lata , Qi Youlin , De Girolami Paola C. , Samore Matthew H. , Anderson Greg , Rasheed J. Kamile , Tenover Fred C.
Published: April 2004

This study analyzed the enzymatic basis and molecular epidemiology of amoxicillin-clavulanate-resistant
Escherichia coli
isolated by the microbiology laboratory of a United States tertiary care hospital. From October 1998 to December 1999, all
E. coli
isolates were screened for ampicillin-sulbactam resistance. Of 283 isolates that tested resistant to ampicillin-sulbactam, 69 unique patient isolates were also resistant to amoxicillin-clavulanate by disk diffusion testing (zone diameter ≤ 13 mm). These amoxicillin-clavulanate-resistant
E. coli
isolates underwent agar dilution testing, pulsed-field gel electrophoresis, PCR analysis, and isoelectric focusing. The mean age of study patients was 52 years; 78% were female. Among the isolates, 12 were nosocomial (rate of amoxicillin-clavulanate resistance = 4.7%) and 57 were community acquired (rate of amoxicillin-clavulanate resistance = 2.8%). No predominant strain was identified. By agar dilution testing, 67 isolates were nonsusceptible (39 resistant and 28 intermediate) to amoxicillin-clavulanate and 37 were piperacillin-tazobactam resistant but only 8 were ceftazidime resistant (ceftazidime MIC ≥ 32 μg/ml). Two isolates were susceptible to amoxicillin-clavulanic acid by agar dilution, although they were resistant by disk diffusion testing. The distribution of β-lactamases was as follows: the TEM type alone was found in 52 isolates, the AmpC type was found in 4 isolates (2 identified as containing CMY-2), the TEM type and CMY-2 were found in 2 isolates, and the OXA type was found in 1 isolate. Also, there was one isolate with the TEM type and the SHV type and one with the TEM type and a second, unidentified enzyme. Among the isolates with TEM-type enzymes, two extended-spectrum β-lactamase-producing isolates were identified but two isolates with inhibitor-resistant TEM (IRT) enzymes (one with TEM-34 [IRT-6] and the other with a novel enzyme [tentatively assigned the designation TEM-122]) were more interesting.
Fungus-Elicited Metabolites from Plants as an Enriched Source for New Leishmanicidal Agents: Antifungal Phenyl-Phenalenone Phytoalexins from the Banana Plant (
Musa acuminata
) Target Mitochondria of
Leishmania donovani
Promastigotes
Authors: Luque-Ortega Juan Román , Martínez Silvia , Saugar José María , Izquierdo Laura R. , Abad Teresa , Luis Javier G. , Piñero José , Valladares Basilio , Rivas Luis
Published: April 2004

Two antifungal phenyl-phenalenone phytoalexins isolated from the banana plant (
Musa acuminata
) elicited with the fungus
Fusarium oxysporum
, together with a methoxy derivative of one of them and two epoxide precursors of their chemical synthesis, were tested for leishmanicidal activity on
Leishmania donovani
promastigotes and
L. infantum
amastigotes. Drugs inhibited proliferation of both forms of the parasite with a 50% lethal concentration range between 10.3 and 68.7 μg/ml. Their lethal mechanism was found linked to the respiratory chain by a systematic approach, including electron microscopy, measurement of the oxygen consumption rate on digitonin-permeabilized promastigotes, and enzymatic assays on a mitochondrial enriched fraction. Whereas the whole set of compounds inhibited the activity of fumarate reductase in the mitochondrial fraction (50% effective concentration [EC
50
] between 33.3 and 78.8 μg/ml) and on purified enzyme (EC
50
= 53.3 to 115 μg/ml), inhibition for succinate dehydrogenase was only observed for the two phytoalexins with the highest leishmanicidal activity: anigorufone and its natural analogue 2-methoxy-9-phenyl-phenalen-1-one (EC
50
= 33.5 and 59.6 μg/ml, respectively). These results provided a new structural motif, phenyl-phenalenone, as a new lead for leishmanicidal activity, and support the use of plant extracts enriched in antifungal phytoalexins, synthesized under fungal challenge, as a more rational and effective strategy to screen for new plant leishmanicidal drugs.
In Vitro Activities of the β-Lactamase Inhibitors Clavulanic Acid, Sulbactam, and Tazobactam Alone or in Combination with β-Lactams against Epidemiologically Characterized Multidrug-Resistant
Acinetobacter baumannii
Strains
Authors: Higgins Paul G. , Wisplinghoff Hilmar , Stefanik Danuta , Seifert Harald
Published: April 2004

Acinetobacter baumannii
is an important nosocomial pathogen usually in the context of serious underlying disease. Multidrug resistance in these organisms is frequent. The β-lactamase inhibitors clavulanic acid, sulbactam, and tazobactam have intrinsic activity against
Acinetobacter
strains. To evaluate their potential therapeutic usefulness, we determined the in vitro activity of ampicillin, sulbactam, ampicillin-sulbactam, cefoperazone, cefoperazone-sulbactam, piperacillin, piperacillin-sulbactam, tazobactam, piperacillin-tazobactam, amoxicillin, clavulanic acid, amoxicillin-clavulanic acid, ticarcillin, and ticarcillin-clavulanic acid against multidrug-resistant
A. baumannii
. All isolates were epidemiologically characterized by RAPD [random(ly) amplified polymorphic DNA] analysis and/or pulsed-field gel electrophoresis and represented different strain types, including sporadic strains, as well as outbreak-related strains. The MICs were determined by agar dilution on Mueller-Hinton agar (using fixed concentrations, as well as fixed ratios for β-lactamase inhibitors) and the E-test. The majority of E-test results were within two dilutions of those recorded by agar dilution, with the exception of piperacillin-tazobactam. Sulbactam was superior to clavulanic acid and tazobactam and may represent an alternative treatment option for infections due to multiresistant
A. baumannii
strains. β-Lactamase inhibitors have intrinsic activity but do not enhance activity of β-lactams against
A. baumannii
. Testing with the inhibitor added at a fixed concentration as recommended for piperacillin-tazobactam and ticarcillin-clavulanic acid by the National Committee for Clinical Laboratory Standards may falsely suggest high activity or gives uninterpretable results due to trailing. If combinations are used for testing, fixed ratios may give more useful results.
Functional Genomic Analysis of Fluconazole Susceptibility in the Pathogenic Yeast
Candida glabrata
: Roles of Calcium Signaling and Mitochondria
Authors: Kaur Rupinder , Castaño Irene , Cormack Brendan P.
Published: April 2004

The pathogenic yeast
Candida glabrata
exhibits innate resistance to fluconazole, the most commonly used antifungal agent. By screening a library of 9,216 random insertion mutants, we identified a set of 27 genes which upon mutation, confer altered fluconazole susceptibility in
C. glabrata
. Homologues of three of these genes have been implicated in azole and/or drug resistance in
Saccharomyces cerevisiae
: two of these belong to the family of ABC transporters (
PDR5
and
PDR16
), and one is involved in retrograde signaling from mitochondria to nucleus (
RTG2
). The remaining 24 genes are involved in diverse cellular functions, including ribosomal biogenesis and mitochondrial function, activation of RNA polymerase II transcription, nuclear ubiquitin ligase function, cell wall biosynthesis, and calcium homeostasis. We characterized two sets of mutants in more detail. Strains defective in a putative plasma membrane calcium channel (Cch1-Mid1) were modestly more susceptible to fluconazole but showed a significant loss of viability upon prolonged fluconazole exposure, suggesting that calcium signaling is required for survival of azole stress in
C. glabrata
. These mutants were defective in calcium uptake in response to fluconazole exposure. The combined results suggest that, in the absence of Ca
2+
signaling, fluconazole has a fungicidal rather than a fungistatic effect on
C. glabrata
. The second set of mutants characterized in detail were defective in mitochondrial assembly and organization, and these exhibited very high levels of fluconazole resistance. Further analysis of these mutants indicated that in
C. glabrata
a mechanism exists for reversible loss of mitochondrial function that does not involve loss of mitochondrial genome and that
C. glabrata
can switch between states of mitochondrial competence and incompetence in response to fluconazole exposure.
Novel Polysulfated Galactose-Derivatized Dendrimers as Binding Antagonists of Human Immunodeficiency Virus Type 1 Infection
Authors: Kensinger Richard D. , Catalone Bradley J. , Krebs Fred C. , Wigdahl Brian , Schengrund Cara-Lynne
Published: April 2004

Evidence indicates that galactosyl ceramide (GalCer) and its 3′-sulfated derivative, sulfatide (SGalCer), may act as alternate coreceptors for human immunodeficiency virus type 1 (HIV-1) in CD4
−
cells. Glycosphingolipids (GSLs) may also be necessary for fusion of HIV-1 and host cell membranes. Using an enzyme-linked immunosorbent assay to determine which GSL was the best ligand for both recombinant and virus-associated gp120, we found that SGalCer was the best ligand for each rgp120 and HIV-1 isolate tested. Therefore, novel multivalent glycodendrimers, which mimic the carbohydrate clustering reportedly found in lipid rafts, were synthesized based on the carbohydrate moiety of SGalCer. Here we describe the synthesis of a polysulfated galactose functionalized, fifth generation DAB dendrimer (PS Gal 64mer), containing on average two sulfate groups per galactose residue. Its ability to inhibit HIV-1 infection of cultured indicator cells was compared to that of dextran sulfate (DxS), a known, potent, binding inhibitor of HIV-1. The results indicate that the PS Gal 64mer inhibited infection by the HIV-1 isolates tested as well as DxS.
Use of Monte Carlo Simulations To Select Therapeutic Doses and Provisional Breakpoints of BAL9141
Authors: Mouton Johan W. , Schmitt-Hoffmann Anne , Shapiro Stuart , Nashed Norman , Punt Nieko C.
Published: April 2004

BAL9141, a new antimicrobial agent belonging to the class of parenteral pyrrolidinone-3-ylidenemethyl cephalosporins, is active against most gram-positive microorganisms, including methicillin-resistant variants (methicillin-resistant
Staphylococcus aureus
[MRSA] and methicillin-resistant
Staphylococcus epidermidis
[MRSE]), as well as against penicillin-resistant pneumococci (PRP) and many gram-negative microorganisms. BAL9141 is administered as the prodrug BAL5788, which is rapidly converted to BAL9141 by plasma esterases. Pharmacokinetic (PK) data obtained in a previous multiple ascending dose study were used to fit a population PK model to using the NPEM2 program, yielding PK parameter estimates and its covariance matrix for BAL9141. These estimates and matrix were used to perform Monte Carlo simulations (MCSs) and obtain unbiased target attainment rates (TARs) for various time periods during which the concentration remains above the MIC (
T
>MIC
). Assuming a
T
>MIC
of 40%, TARs of 100% were reached with a dose of 500 mg/liter every 12 h for pathogens with MICs of 2 mg/liter and with a dose of 750 mg/liter every 12 h for pathogens with MICs of 4 mg/liter. Because MICs are ≤2 mg/liter for most strains of MRSA, MRSE, and PRP (with some strains showing an MIC of 4 mg/liter), a dosing regimen of 750 mg every 12 h is proposed for clinical studies. The corresponding provisional breakpoint is
S
(susceptible) ≤ 4 mg/liter.
Mouse Model of Cervicovaginal Toxicity and Inflammation for Preclinical Evaluation of Topical Vaginal Microbicides
Authors: Catalone Bradley J. , Kish-Catalone Tina M. , Budgeon Lynn R. , Neely Elizabeth B. , Ferguson Maelee , Krebs Fred C. , Howett Mary K. , Labib Mohamed , Rando Robert , Wigdahl Brian
Published: April 2004

Clinical trials evaluating the efficacy of nonoxynol-9 (N-9) as a topical microbicide concluded that N-9 offers no in vivo protection against human immunodeficiency virus type 1 (HIV-1) infection, despite demonstrated in vitro inactivation of HIV-1 by N-9. These trials emphasize the need for better model systems to determine candidate microbicide effectiveness and safety in a preclinical setting. To that end, time-dependent in vitro cytotoxicity, as well as in vivo toxicity and inflammation, associated with N-9 exposure were characterized with the goal of validating a mouse model of microbicide toxicity. In vitro studies using submerged cell cultures indicated that human cervical epithelial cells were inherently more sensitive to N-9-mediated damage than human vaginal epithelial cells. These results correlated with in vivo findings obtained by using Swiss Webster mice in which intravaginal inoculation of 1% N-9 or Conceptrol gel (containing 4% N-9) resulted in selective and acute disruption of the cervical columnar epithelial cells 2 h postapplication accompanied by intense inflammatory infiltrates within the lamina propria. Although damage to the cervical epithelium was apparent out to 8 h postapplication, these tissues resembled control tissue by 24 h postapplication. In contrast, minimal damage and infiltration were associated with both short- and long-term exposure of the vaginal mucosa to either N-9 or Conceptrol. These analyses were extended to examine the relative toxicity of polyethylene hexamethylene biguanide (PEHMB), a polybiguanide compound under evaluation as a candidate topical microbicide. In similar studies, in vivo exposure to 1% PEHMB caused minimal damage and inflammation of the genital mucosa, a finding consistent with the demonstration that PEHMB was >350-fold less cytotoxic than N-9 in vitro. Collectively, these studies highlight the murine model of toxicity as a valuable tool for the preclinical assessment of toxicity and inflammation associated with exposure to candidate topical microbicides.
Protecting rockfish through gear design: development of a selective flatfish trawl for the U.S. west coast bottom trawl fishery
Authors: King Sarah E , Hannah Robert W , Parker Steven J , Matteson Keith M , Berkeley Steven A
Published: April 2004

This study evaluated the potential to reduce the bycatch of rockfish (Sebastes spp.) in a flatfish trawl fishery by developing a selective trawl that separates flatfish from rockfish using differences in their behavior as the trawl net approaches. Using an alternate haul, randomized block design, we compared catches between a commonly used combination trawl and a newly designed, low-rise trawl incorporating a "cutback" headrope. For most of the target flatfish species, the experimental trawl had a 2559% higher catch rate (kilograms) than the control net (P < 0.05), consistent with greater footrope length in the experimental trawl. Increased catches of some small rockfish, skates, and sablefish (Anoplopoma fimbria) were also observed (P < 0.05). The catches of many larger roundfish and rockfish species, including canary rockfish (Sebastes pinniger), redstripe rockfish (Sebastes proriger), Pacific hake (Merluccius productus), and shortspine thornyhead (Sebastolobus alascanus) more than 25 cm long, were reduced from 34 to 97% in the experimental trawl (P < 0.05). Under weak-stock management conditions, in which the bycatch of overfished rockfish species limits directed flatfish fisheries, trawl designs that permit nontarget species to escape before entrainment can significantly reduce bycatch mortality and prevent premature closures of directed flatfish fisheries.
Size structural change in lightly exploited coral reef fish communities: evidence for weak indirect effects
Authors: Dulvy N K , Polunin N VC , Mill A C , Graham N AJ
Published: April 2004

Exploitation influences community structure directly through preferential removal of larger-bodied fishes and indirectly because larger-bodied fishes may exert top-down control upon other community members. We consider the direct and indirect effects of subsistence exploitation upon the size structure of coral reef fish communities along an island-scale spatial gradient of fishing intensity. We explored the effect of fishing intensity and sample date (three dates over a year) at six islands and the overall effect of fishing intensity averaged over sample dates at 13 islands. Fishing intensity consistently explained more of the variation in the size metrics than sample date. In response to exploitation, the mean weight of individuals declined by 52%, the weighted average maximum size (Lmax) declined by 25%, and slopes of community size spectra steepened. The larger size classes (>26 cm) declined in relative numbers by 63% and relative biomass by 61% in response to exploitation. However, the numbers and biomass of the three smallest size classes (<25 cm) increased by 31% and 9%, respectively, in response to exploitation. This increased abundance is consistent with a weak compensatory response presumably from a reduction in predation upon smaller size classes as an indirect effect of exploiting larger size classes.
Crude-oil-degrading thermophilic bacterium isolated from an oil field
Authors: Hao Ruixia , Lu Anhuai , Wang Guanyu
Published: April 2004

Thermophilic bacterium strain C2, which has the ability to transform crude oils, was isolated from the reservoir of the Shengli oil field in East China. The Gram-negative, rod-shaped, nonmotile cells were grown at a high temperature, up to 83 °C, in the neutral to alkaline pH range. Depending on the culture conditions, the organism occurred as single rods or as filamentous aggregates. Strain C2 was grown chemoorganotrophically and produced metabolites, such as volatile fatty acids, 1,2-benzenedicarboxylic acid, bis(2-ethylhexyl)ester, dibutyl phthalate, and di-n-octyl phthalate. It could metabolize different organic substrates (acetate, D-glucose, fructose, glycerol, maltose, pyruvate, starch, sucrose, xylose, hexadecane). The G+C content (68 mol%) and the 16S rRNA sequence of strain C2 indicated that the isolate belonged to the genus Thermus. The strain affected different crude oils and changed their physical and chemical properties. The biochemical interactions between crude oils and strain C2 follow distinct trends characterized by a group of chemical markers (saturates, aromatics, resins, asphaltenes). Those trends show an increase in saturates and a decrease in aromatics, resins, and asphaltenes. The bioconversion of crude oils leads to an enrichment in lighter hydrocarbons and an overall redistribution of these hydrocarbons.Key words: thermophile, metabolite, crude oil, degradation, conversion.
Flatworm nerve–muscle: structural and functional analysis
Authors: Halton D W , Maule A G
Published: April 2004

Platyhelminthes occupy a unique position in nerve–muscle evolution, being the most primitive of metazoan phyla. Essentially, their nervous system consists of an archaic brain and associated pairs of longitudinal nerve cords cross-linked as an orthogon by transverse commissures. Confocal imaging reveals that these central nervous system elements are in continuity with an array of peripheral nerve plexuses which innervate a well-differentiated grid work of somatic muscle as well as a complexity of myofibres associated with organs of attachment, feeding, and reproduction. Electrophysiological studies of flatworm muscles have exposed a diversity of voltage-activated ion channels that influence muscle contractile events. Neuronal cell types are mainly multi- and bi-polar and highly secretory in nature, producing a heterogeneity of vesicular inclusions whose contents have been identified cytochemically to include all three major types of cholinergic, aminergic, and peptidergic messenger molecules. A landmark discovery in flatworm neuro biology was the biochemical isolation and amino acid sequencing of two groups of native neuropeptides: neuro peptide F and FMRFamide-related peptides (FaRPs). Both families of neuropeptide are abundant and broadly distributed in platyhelminths, occurring in neuronal vesicles in representatives of all major flatworm taxa. Dual localization studies have revealed that peptidergic and cholinergic substances occupy neuronal sets separate from those of serotoninergic components. The physiological actions of neuronal messengers in flatworms are beginning to be established, and where examined, FaRPs and 5-HT are myoexcitatory, while cholinomimetic substances are generally inhibitory. There is immunocytochemical evidence that FaRPs and 5-HT have a regulatory role in the mechanism of egg assembly. Use of muscle strips and (or) muscle fibres from free-living and parasitic flatworms has provided baseline information to indicate that muscle responses to FaRPs are mediated by a G-protein-coupled receptor, and that the signal transduction pathway for contraction involves the second messengers cAMP and protein kinase C.
A SURVEY ON ALGORITHMIC ASPECTS OF TANDEM REPEATS EVOLUTION
Authors: RIVALS ERIC
Published: April 2004

Local repetitions in genomes are called tandem repeats. A tandem repeat contains multiple, but slightly different copies of a repeated unit. It changes over time as the copies are altered by mutations, when additional copies are created by amplification of an existing copy, or when a copy is removed by contraction. Theses changes let tandem repeats evolve dynamically. From this statement follow two problems. TANDEM REPEAT HISTORY aims at recovering the history of amplifications and mutations that produced the tandem repeat sequence given as input. Given the tandem repeat sequences at the same genomic location in two individuals and a cost function for amplifications, contractions, and mutations, the purpose of TANDEM REPEAT ALLELE ALIGNMENT is to find an alignment of the sequences having minimal cost. We present a survey of these two problems that allow to investigate evolutionary mechanisms at work in tandem repeats.
WORDS GUARANTEEING MINIMUM IMAGE
Authors: MARGOLIS S. W. , PIN J.-E. , VOLKOV M. V.
Published: April 2004

Given a positive integer n and a finite alphabet Σ, a word w over Σ is said to guarantee minimum image if, for every homomorphism φ from the free monoid Σ* over Σ into the monoid of all transformations of an n-element set, the range of the transformation wφ has the minimum cardinality among the ranges of all transformations of the form vφ where v runs over Σ*. Although the existence of words guaranteeing minimum image is pretty obvious, the problem of their explicit description is very far from being trivial. Sauer and Stone in 1991 gave a recursive construction for such a word w but the length of their word was doubly exponential (as a function of n). We first show that some known results of automata theory immediately lead to an alternative construction that yields a simpler word that guarantees minimum image: it has exponential length, more precisely, its length is O(|Σ|⅙(n3-n)). Then with some more effort, we find a word guaranteeing minimum image similar to that of Sauer and Stone but of length O(|Σ|½(n2-n)). On the other hand, we prove that the length of any word guaranteeing minimum image cannot be less than |Σ|n-1.
MATRIX INDICATORS FOR SUBWORD OCCURRENCES AND AMBIGUITY
Authors: MATEESCU ALEXANDRU , SALOMAA ARTO
Published: April 2004

The paper investigates inequalities between the numbers of different (scattered) subword occurrences. The Parikh matrix recently introduced is an efficient tool. We give various characterizations for Parikh matrices. Of special interest is the case where the matrix determines the word uniquely. We investigate such matrix unambiguous words. The considerations are extended to concern languages. Several open problems and problem areas are indicated.
CONJECTURES AND RESULTS ON MORPHISMS GENERATING k-POWER-FREE WORDS
Authors: RICHOMME GWÉNAËL , SÉÉBOLD PATRICE
Published: April 2004

After giving an overview about morphisms and k-power-freeness (k∈ IN ), we present several conjectures, and partially solve one of those by giving, for any integer k≥3, binary morphisms which are not k-power-free, but which are (k+1)-power-free and generate k-power-free infinite words.
DYNAMICAL DETAILED BALANCE AND LOCAL KMS CONDITION FOR NON-EQUILIBRIUM STATES
Authors: ACCARDI LUIGI , IMAFUKU KENTARO
Published: April 2004

The principle of detailed balance is at the basis of equilibrium physics and is equivalent to the Kubo–Martin–Schwinger (KMS) condition (under quite general assumptions). In the present paper we prove that a large class of non-equilibrium quantum systems satisfies a dynamical generalization of the detailed balance condition (dynamical detailed balance) expressing the fact that all the micro-currents, associated to the Bohr frequencies are constant. The usual (equilibrium) detailed balance condition is characterized by the property that this constant is identically zero. From this we deduce a simple and experimentally measurable relation expressing the microcurrent associated to a transition between two levels ∊m→∊n as a linear combination of the occupation probabilities of the two levels, with coefficients given by the generalized susceptivities (transport coefficients). We then give a second characterization of the dynamical detailed balance condition using a master equation rather than the microcurrents. Finally we show that these two conditions are equivalent to a "local" generalization of the usual KMS condition. Summing up: rather than postulating some ansatz on the basis of phenomenological models or of numerical simulations, we deduce, directly in the quantum domain and from fundamental principles, some natural and simple non equilibrium generalizations of the three main characterizations of equilibrium states. Then we prove that these three, apparently very far, conditions are equivalent. These facts support our convinction that these three equivalent conditions capture a universal aspect of non equilibrium phenomena.
FROM THE MATHEMATICAL KINETIC THEORY TO MODELLING COMPLEX SYSTEMS IN APPLIED SCIENCES
Authors: BELLOMO NICOLA
Published: April 2004

This paper deals with the design of a nonequilibrium statistical mechanics theory developed to model large systems of interacting individuals. Interactions being ruled, not only by laws of classical mechanics, but also by some intelligent or organized behaviour. The paper technically shows, also by reasoning on specific examples, how the theory can be applied to model large complex systems in natural and applied sciences.
ANALOGICAL MODEL INFERRED FROM TIME DOMAIN ANALYSIS AND GENERATING ORGAN PIPES SELF SUSTAINED-TONE
Authors: BOTTIGLIERI MILENA , DE LAURO ENZA , DE MARTINO SALVATORE , FALANGA MARIA ROSARIA
Published: April 2004

We have recorded acoustic signals emitted by organ pipes in a variety of experimental framework. We have analyzed the recorded sounds with nonlinear techniques in time domain. Starting from this analysis we have extracted their relevant characteristics. Finally, we have constructed simple and suitable analogical model, able to reproduce the registered waveform and sound in listening.