bluuebunny/crossref_metadata_2025 huggingface dataset:
Optimal Static Characteristics of Amphibious PAR Vehicles
Authors: Matveev Konstantin I.
Published: July 2008

Power augmented ram platform vehicle is a high-performance air-assisted amphibious transport of the new generation. The objective of this study is to determine the optimal position of the platform flap for low-speed operations on water and solid ground. The static friction of vehicle hulls on solid surfaces and the static deformation of the water surface are combined with results of the ideal jet theory to predict forces and the platform position. The optimal values for the flap gap are obtained that maximize the recovered static thrust on solid surfaces and the vehicle vertical position on the water surface.
Putative Virulence Traits and Pathogenicity of
Vibrio cholerae
Non-O1, Non-O139 Isolates from Surface Waters in Kolkata, India
Authors: Bag Prasanta K. , Bhowmik Poulami , Hajra Tapas K. , Ramamurthy T. , Sarkar Pradipta , Majumder Mrinmoyee , Chowdhury Goutam , Das Suresh C.
Published: July 2008

Vibrio cholerae
non-O1, non-O139 was isolated from natural surface waters from different sites sampled in diarrhea endemic zones in Kolkata, India. Twenty-one of these isolates were randomly selected and included in the characterization. The multiserogroup isolates were compared by their virulence traits with a group of clinical non-O1, non-O139 isolates from the same geographic area. Of the 21 environmental isolates, 6 and 14 strains belonged to Heiberg groups I and II, respectively. Three of the environmental isolates showed resistance to 2,2-diamine-6,7-diisopropylpteridine phosphate. All of the non-O1, non-O139 strains were positive for
toxR
, and except for one environmental isolate, none of them were positive for
tcpA
in the PCR assay. None of the isolates were positive for genes encoding cholera toxin (
ctxA
), heat-stable toxin (
est
), heat-labile toxin (
elt
), and Shiga toxin variants (
stx
) of
Escherichia coli
. Additionally, except for one environmental isolate (PC32), all were positive for the gene encoding El Tor hemolysin (
hly
). The culture supernatants of 86% (18 of 21) of the environmental isolates showed a distinct cytotoxic effect on HeLa cells, and some of these strains also produced cell-rounding factor. The lipase, protease, and cell-associated hemagglutination activities and serum resistance properties of the environmental and clinical isolates did not differ much. However, seven environmental isolates exhibited very high hemolytic activities (80 to 100%), while none of the clinical strains belonged to this group. The environmental isolates manifested three adherence patterns, namely, carpet-like, diffuse, and aggregative adherence, and the clinical isolates showed diffuse adherence on HeLa cells. Of the 11 environmental isolates tested for enteropathogenic potential, 8 (73%) induced positive fluid accumulation (≥100) in a mouse model, and the reactivities of these isolates were comparable to those of clinical strains of non-O1, non-O139 and toxigenic O139
V. cholerae
. Comparison of the counts of the colonized environmental and clinical strains in the mouse intestine showed that the organisms of both groups had similar colonizing efficiencies. These findings indicate the presence of potentially pathogenic
V. cholerae
non-O1, non-O139 strains in surface waters of the studied sites in Kolkata.
Cloning and Overexpression of Alkaline Phosphatase PhoK from
Sphingomonas
sp. Strain BSAR-1 for Bioprecipitation of Uranium from Alkaline Solutions
Authors: Nilgiriwala Kayzad S. , Alahari Anuradha , Rao Amara Sambasiva , Apte Shree Kumar
Published: July 2008

Cells of
Sphingomonas
sp. strain BSAR-1 constitutively expressed an alkaline phosphatase, which was also secreted in the extracellular medium. A null mutant lacking this alkaline phosphatase activity was isolated by Tn
5
random mutagenesis. The corresponding gene, designated
phoK
, was cloned and overexpressed in
Escherichia coli
strain BL21(DE3). The resultant
E. coli
strain EK4 overexpressed cellular activity 55 times higher and secreted extracellular PhoK activity 13 times higher than did BSAR-1. The recombinant strain very rapidly precipitated >90% of input uranium in less than 2 h from alkaline solutions (pH, 9 ± 0.2) containing 0.5 to 5 mM of uranyl carbonate, compared to BSAR-1, which precipitated uranium in >7 h. In both strains BSAR-1 and EK4, precipitated uranium remained cell bound. The EK4 cells exhibited a much higher loading capacity of 3.8 g U/g dry weight in <2 h compared to only 1.5 g U/g dry weight in >7 h in BSAR-1. The data demonstrate the potential utility of genetically engineering PhoK for the bioprecipitation of uranium from alkaline solutions.
Genetic Diversity of
Porphyromonas gingivalis
Isolates Recovered from Single “Refractory” Periodontitis Sites
Authors: Enersen Morten , Olsen Ingar , Caugant Dominique A.
Published: July 2008

Multilocus sequence typing and
fimA
genotyping were performed on
Porphyromonas gingivalis
isolates from 15 subjects with “refractory” periodontitis. Several sequence types were detected for most individual pockets. The variation indicated recombination at the
recA
and
pepO
genes. The prevalence of
fimA
genotypes II and IV confirmed their association with periodontitis.
Prevalence and Genetic Characterization of
Cryptosporidium
Isolates from Common Brushtail Possums (
Trichosurus vulpecula
) Adapted to Urban Settings
Authors: Hill Nichola J. , Deane Elizabeth M. , Power Michelle L.
Published: July 2008

The common brushtail possum (
Trichosurus vulpecula
) is one of the most abundant native marsupials in urban Australia, having successfully adapted to utilize anthropogenic resources. The habituation of possums to food and shelter available in human settlements has facilitated interaction with people, pets, and zoo animals, increasing the potential for transmission of zoonotic
Cryptosporidium
pathogens. This study sought to examine the identity and prevalence of
Cryptosporidium
species occurring in possums adapted to urban settings compared to possums inhabiting remote woodlands far from urban areas and to characterize the health of the host in response to oocyst shedding. Findings indicated that both populations were shedding oocysts of the same genotype (brushtail possum 1 [BTP1]) that were genetically and morphologically distinct from zoonotic species and genotypes and most closely related to
Cryptosporidium
species from marsupials. The urban population was shedding an additional five
Cryptosporidium
isolates that were genetically distinct from BTP1 and formed a sister clade with
Cryptosporidium parvum
and
Cryptosporidium hominis
. Possums that were shedding oocysts showed no evidence of pathogenic changes, including elevated levels of white blood cells, diminished body condition (body mass divided by skeletal body length), or reduced nutritional state, suggesting a stable host-parasite relationship typical of
Cryptosporidium
species that are adapted to the host. Overall,
Cryptosporidium
occurred with a higher prevalence in possums from urban habitat (11.3%) than in possums from woodland habitat (5.6%); however, the host-specific nature of the genotypes may limit spillover infection in the urban setting. This study determined that the coexistence of possums with sympatric populations of humans, pets, and zoo animals in the urban Australian environment is unlikely to present a threat to public health safety.
Relative Importance of H
2
and H
2
S as Energy Sources for Primary Production in Geothermal Springs
Authors: D'Imperio Seth , Lehr Corinne R. , Oduro Harry , Druschel Greg , Kühl Michael , McDermott Timothy R.
Published: July 2008

Geothermal waters contain numerous potential electron donors capable of supporting chemolithotrophy-based primary production. Thermodynamic predictions of energy yields for specific electron donor and acceptor pairs in such systems are available, although direct assessments of these predictions are rare. This study assessed the relative importance of dissolved H
2
and H
2
S as energy sources for the support of chemolithotrophic metabolism in an acidic geothermal spring in Yellowstone National Park. H
2
S and H
2
concentration gradients were observed in the outflow channel, and vertical H
2
S and O
2
gradients were evident within the microbial mat. H
2
S levels and microbial consumption rates were approximately three orders of magnitude greater than those of H
2
.
Hydrogenobaculum
-like organisms dominated the bacterial component of the microbial community, and isolates representing three distinct 16S rRNA gene phylotypes (phylotype = 100% identity) were isolated and characterized. Within a phylotype, O
2
requirements varied, as did energy source utilization: some isolates could grow only with H
2
S, some only with H
2
, while others could utilize either as an energy source. These metabolic phenotypes were consistent with in situ geochemical conditions measured using aqueous chemical analysis and in-field measurements made by using gas chromatography and microelectrodes. Pure-culture experiments with an isolate that could utilize H
2
S and H
2
and that represented the dominant phylotype (70% of the PCR clones) showed that H
2
S and H
2
were used simultaneously, without evidence of induction or catabolite repression, and at relative rate differences comparable to those measured in ex situ field assays. Under in situ-relevant concentrations, growth of this isolate with H
2
S was better than that with H
2
. The major conclusions drawn from this study are that phylogeny may not necessarily be reliable for predicting physiology and that H
2
S can dominate over H
2
as an energy source in terms of availability, apparent in situ consumption rates, and growth-supporting energy.
Prevalence and Genetic Characterization of Shiga Toxin-Producing
Escherichia coli
Isolates from Slaughtered Animals in Bangladesh
Authors: Islam Mohammad A. , Mondol Abdus S. , de Boer Enne , Beumer Rijkelt R. , Zwietering Marcel H. , Talukder Kaisar A. , Heuvelink Annet E.
Published: July 2008

To determine the prevalence of Shiga toxin (Stx)-producing
Escherichia coli
(STEC) in slaughter animals in Dhaka, Bangladesh, we collected rectal contents immediately after animals were slaughtered. Of the samples collected from buffalo (
n
= 174), cows (
n
= 139), and goats (
n
= 110), 82.2%, 72.7%, and 11.8% tested positive for
stx
1
and/or
stx
2
, respectively. STEC could be isolated from 37.9%, 20.1%, and 10.0% of the buffalo, cows, and goats, respectively. STEC O157 samples were isolated from 14.4% of the buffalo, 7.2% of the cows, and 9.1% of the goats. More than 93% (
n
= 42) of the STEC O157 isolates were positive for the
stx
2
,
eae
,
katP
,
etpD
, and enterohemorrhagic
E. coli hly
(
hly
EHEC
) virulence genes. STEC O157 isolates were characterized by seven recognized phage types, of which types 14 (24.4%) and 31 (24.4%) were predominant. Subtyping of the 45 STEC O157 isolates by pulsed-field gel electrophoresis showed 37 distinct restriction patterns, suggesting a heterogeneous clonal diversity. In addition to STEC O157, 71 STEC non-O157 strains were isolated from 60
stx
-positive samples from 23.6% of the buffalo, 12.9% of the cows, and 0.9% of the goats. The STEC non-O157 isolates belonged to 36 different O groups and 52 O:H serotypes. Unlike STEC O157, most of the STEC non-O157 isolates (78.9%) were positive for
stx
1
. Only 7.0% (
n
= 5) of the isolates were positive for
hly
EHEC
, and none was positive for
eae
,
katP
, and
etpD
. None of the isolates was positive for the
iha
,
toxB
, and
efa1
putative adhesion genes. However, 35.2% (
n
= 25), 11.3% (
n
= 8), 12.7% (
n
= 9), and 12.7% (
n
= 9) of the isolates were positive for the
lpf
O113
,
saa
,
lpfA
O157/01-141
, and
lpfA
O157/OI-154
genes, respectively. The results of this study provide the first evidence that slaughtered animals like buffalo, cows, and goats in Bangladesh are reservoirs for STEC, including the potentially virulent STEC strain O157.
Global Gene Expression Profiles Suggest an Important Role for Nutrient Acquisition in Early Pathogenesis in a Plant Model ofPseudomonas aeruginosaInfection
Authors: Weir Tiffany L. , Stull Valerie J. , Badri Dayakar , Trunck Lily A. , Schweizer Herbert P. , Vivanco Jorge
Published: July 2008

AlthoughPseudomonas aeruginosais an opportunistic pathogen that does not often naturally infect alternate hosts, such as plants, the plant-P. aeruginosamodel has become a widely recognized system for identifying new virulence determinants and studying the pathogenesis of the organism. Here, we examine how both host factors andP. aeruginosaPAO1 gene expression are affected in planta after infiltration into incompatible and compatible cultivars of tobacco (Nicotiana tabacumL.).N. tabacumhas a resistance gene (N) against tobacco mosaic virus, and although resistance to PAO1 infection is correlated with the presence of a dominant N gene, our data suggest that it is not a factor in resistance against PAO1. We did observe that the resistant tobacco cultivar had higher basal levels of salicylic acid and a stronger salicylic acid response upon infiltration of PAO1. Salicylic acid acts as a signal to activate defense responses in plants, limiting the spread of the pathogen and preventing access to nutrients. It has also been shown to have direct virulence-modulating effects onP. aeruginosa. We also examined host effects on the pathogen by analyzing global gene expression profiles of bacteria removed from the intracellular fluid of the two plant hosts. We discovered that the availability of micronutrients, particularly sulfate and phosphates, is important for in planta pathogenesis and that the amounts of these nutrients made available to the bacteria may in turn have an effect on virulence gene expression. Indeed, there are several reports suggesting thatP. aeruginosavirulence is influenced in mammalian hosts by the availability of micronutrients, such as iron and nitrogen, and by levels of O2.
Microscopic and Molecular Studies of the Diversity of Free-Living Protozoa in Meat-Cutting Plants
Authors: Vaerewijck Mario J. M. , Sabbe Koen , Baré Julie , Houf Kurt
Published: July 2008

The diversity of free-living protozoa in five meat-cutting plants was determined. Light microscopy after enrichment culturing was combined with sequencing of PCR-amplified, denaturing gradient gel electrophoresis (DGGE)-separated 18S rRNA gene fragments, which was used as a fast screening method. The general results of the survey showed that a protozoan community of amoebae, ciliates, and flagellates was present in all of the plants. Protozoa were detected mainly in floor drains, in standing water on the floor, on soiled bars of cutting tables, on plastic pallets, and in out-of-use hot water knife sanitizers, but they were also detected on surfaces which come into direct contact with meat, such as conveyer belts, working surfaces of cutting tables, and needles of a meat tenderizer. After 7 days of incubation at refrigerator temperature, protozoa were detected in about one-half of the enrichment cultures. Based on microscopic observations, 61 morphospecies were found, and
Bodo saltans, Bodo
spp.,
Epistylis
spp.,
Glaucoma scintillans, Petalomonas
spp.,
Prodiscophrya collini
, and
Vannella
sp. were the most frequently encountered identified organisms. Sequencing of DGGE bands resulted in identification of a total of 49 phylotypes, including representatives of the Amoebozoa, Chromalveolata, Excavata, Opisthokonta, and Rhizaria. Sequences of small heterotrophic flagellates were affiliated mainly with the Alveolata (Apicomplexa), Stramenopiles (Chrysophyceae), and Rhizaria (Cercozoa). This survey showed that there is high protozoan species richness in meat-cutting plants and that the species included species related to known hosts of food-borne pathogens.
Self-Cloning Baker's Yeasts That Accumulate Proline Enhance Freeze Tolerance in Doughs
Authors: Kaino Tomohiro , Tateiwa Tetsuya , Mizukami-Murata Satomi , Shima Jun , Takagi Hiroshi
Published: July 2008

We constructed self-cloning diploid baker's yeast strains by disrupting
PUT1
, encoding proline oxidase, and replacing the wild-type
PRO1
, encoding γ-glutamyl kinase, with a
pro1
(
D154N
) or
pro1
(
I150T
) allele. The resultant strains accumulated intracellular proline and retained higher-level fermentation abilities in the frozen doughs than the wild-type strain. These results suggest that proline-accumulating baker's yeast is suitable for frozen-dough baking.
Plant Lesions Promote the Rapid Multiplication of
Escherichia coli
O157:H7 on Postharvest Lettuce
Authors: Brandl M. T.
Published: July 2008

Several outbreaks of
Escherichia coli
O157:H7 infections have been associated with minimally processed leafy vegetables in the United States. Harvesting and processing cause plant tissue damage. In order to assess the role of plant tissue damage in the contamination of leafy greens with
E. coli
O157:H7, the effect of mechanical, physiological, and plant disease-induced lesions on the growth of this pathogen on postharvest romaine lettuce was investigated. Within only 4 h after inoculation, the population sizes of
E. coli
O157:H7 increased 4.0-, 4.5-, and 11.0-fold on lettuce leaves that were mechanically bruised, cut into large pieces, and shredded into multiple pieces, respectively. During the same time,
E. coli
O157:H7 population sizes increased only twofold on leaves that were left intact after harvest. Also, the population size of
E. coli
O157:H7 was 27 times greater on young leaves affected by soft rot due to infection by
Erwinia chrysanthemi
than on healthy middle-aged leaves. Confocal microscopy revealed that leaf tip burn lesions, which are caused by a common physiological disorder of lettuce, harbored dense populations of
E. coli
O157:H7 cells both internally and externally. Investigation of the colonization of cut lettuce stems by
E. coli
O157:H7 showed that the pathogen grew 11-fold over 4 h of incubation after its inoculation onto the stems, from which large amounts of latex were released. The results of this study indicate that plant tissue damage of various types can promote significant multiplication of
E. coli
O157:H7 over a short time and suggest that harvesting and processing are critical control points in the prevention or reduction of
E. coli
O157:H7 contamination of lettuce.
Diversity of Bacterial Endosymbionts of Environmental
Acanthamoeba
Isolates
Authors: Schmitz-Esser Stephan , Toenshoff Elena R. , Haider Susanne , Heinz Eva , Hoenninger Verena M. , Wagner Michael , Horn Matthias
Published: July 2008

Free-living amoebae are frequent hosts for bacterial endosymbionts. In this study, the symbionts of eight novel environmental
Acanthamoeba
strains isolated from different locations worldwide were characterized. Phylogenetic analysis revealed that they were related to one of four evolutionary lineages of amoeba symbionts recognized previously. This study provides evidence for the existence of only a small number of phylogenetically well-separated groups of obligate intracellular endosymbionts of acanthamoebae with global distribution.
Transmission of
Yersinia pseudotuberculosis
in the Pork Production Chain from Farm to Slaughterhouse
Authors: Laukkanen Riikka , Martínez Pilar Ortiz , Siekkinen Kirsi-Maarit , Ranta Jukka , Maijala Riitta , Korkeala Hannu
Published: July 2008

The transmission of
Yersinia pseudotuberculosis
in the pork production chain was followed from farm to slaughterhouse by studying the same 364 pigs from different production systems at farm and slaughterhouse levels. In all, 1,785 samples were collected, and the isolated
Y. pseudotuberculosis
strains were analyzed by pulsed-field gel electrophoresis. The results of microbial sampling were combined with data from an on-farm observation and questionnaire study to elucidate the associations between farm factors and the prevalence of
Y. pseudotuberculosis
. Following the same pigs in the production chain from farm to slaughterhouse, we were able to show similar
Y. pseudotuberculosis
genotypes in live animals, pluck sets (containing tongue, tonsils, esophagus, trachea, heart, lungs, diaphragm, liver, and kidneys), and carcasses and to conclude that
Y. pseudotuberculosis
contamination originates from the farms, is transported to slaughterhouses with pigs, and transfers to pluck sets and carcasses in the slaughter process. The study also showed that the high prevalence of
Y. pseudotuberculosis
in live pigs predisposes carcasses and pluck sets to contamination. When production types and capacities were compared, the prevalence of
Y. pseudotuberculosis
was higher in organic production than in conventional production and on conventional farms with high rather than low production capacity. We were also able to associate specific farm factors with the prevalence of
Y. pseudotuberculosis
by using a questionnaire and on-farm observations. On farms, contact with pest animals and the outside environment and a rise in the number of pigs on the farm appear to increase the prevalence of
Y. pseudotuberculosis
.
Dynamics of Introduced Populations of
Phragmidium violaceum
and Implications for Biological Control of European Blackberry in Australia
Authors: Gomez D. R. , Evans K. J. , Baker J. , Harvey P. R. , Scott E. S.
Published: July 2008

Phragmidium violaceum
causes leaf rust on the European blackberry (
Rubus fruticosus
L. aggregate). Multiple strains of this pathogen have been introduced into southern Australia for the biological control of at least 15 taxa of European blackberry, a nonindigenous, invasive plant. In climates conducive to leaf rust, the intensity of disease varies within and among infestations of the genetically variable host. Genetic markers developed from the selective amplification of microsatellite polymorphic loci were used to assess the population genetic structure and reproductive biology of
P. violaceum
within and among four geographically isolated and diseased infestations of the European blackberry in Victoria, Australia. Despite the potential for long-distance aerial dispersal of urediniospores, there was significant genetic differentiation among all populations, which was not associated with geographic separation. An assessment of multilocus linkage disequilibrium revealed temporal and geographic variation in the occurrence of random mating among the four populations. The presence of sexual spore states and the results of genetic analyses indicated that recombination, and potentially random migration and genetic drift, played an important role in maintaining genotypic variation within populations. Recombination and genetic differentiation in
P. violaceum
, as well as the potential for metapopulation structure, suggest the need to release additional, genetically diverse strains of the biocontrol agent at numerous sites across the distribution of the Australian blackberry infestation for maximum establishment and persistence.
TolC-Dependent Exclusion of Porphyrins in
Escherichia coli
Authors: Tatsumi Ryoko , Wachi Masaaki
Published: July 2008

We found that
Escherichia coli tolC
mutants showed increased sensitivity to 5-aminolevulinic acid (ALA), a precursor of porphyrins. The
tolC
mutant cells grown in the presence of ALA showed a reddish brown color under visible light and a strong red fluorescence under near-UV irradiation. Fluorescence spectrometry and high-performance liquid chromatography analysis showed that the
tolC
mutant cells grown in the presence of ALA accumulated a large amount of coproporphyrin(ogen) intracellularly. In contrast, the wild-type cells produced coproporphyrin extracellularly. The
tolC
mutant cells grown in the presence of ALA, which were capable of surviving in the dark, were killed by near-UV irradiation, suggesting that the intracellular coproporphyrin(ogen) renders these cells photosensitive. These results suggest that the TolC-dependent efflux system is involved in the exclusion of porphyrin(ogen)s in
E. coli
.
Novel Inner Membrane Retention Signals inPseudomonas aeruginosaLipoproteins
Authors: Lewenza Shawn , Mhlanga Musa M. , Pugsley Anthony P.
Published: July 2008

The ultimate membrane localization and function of most of the 185 predictedPseudomonas aeruginosaPAO1 lipoproteins remain unknown. We constructed a fluorescent lipoprotein, CSFPOmlA-ChFP, by fusing the signal peptide and the first four amino acids of theP. aeruginosaouter membrane lipoprotein OmlA to the monomeric red fluorescent protein mCherry (ChFP). When cells were plasmolyzed with 0.5 M NaCl, the inner membrane separated from the outer membrane and formed plasmolysis bays. This permits the direct observation of fluorescence in either the outer or inner membrane. CSFPOmlA-ChFP was shown to localize in the outer membrane by fluorescence microscopy and immunoblotting analysis of inner and outer membrane fractions. The site-directed substitution of the amino acids at positions +2, +3, and +4 in CSFPOmlA-ChFP was performed to test the effects on lipoprotein localization of a series of amino acid sequences selected from a panel of predicted lipoproteins. We confirmed Asp+2and Lys+3Ser+4function as inner membrane retention signals and identified four novel inner membrane retention signals: CK+2V+3E+4, CG+2G+3G+4, CG+2D+3D+4, and CQ+2G+3S+4. These inner membrane retention signals are found in 5% of the 185 predictedP. aeruginosalipoproteins. Full-length chimeras of predicted lipoproteins PA4370 and PA3262 fused to mCherry were shown to reside in the inner membrane and showed a nonuniform or patchy distribution in the membrane. The optical sectioning of cells producing PA4370CGDD-ChFP and PA3262CDSQ-ChFP by confocal microscopy improved the resolution and indicated a helix-like localization pattern in the inner membrane. The method described here permits the in situ visualization of lipoprotein localization and should work equally well for other membrane-associated proteins.
Genomic Analysis of the Role of RNase R in the Turnover ofPseudomonas putidamRNAs
Authors: Fonseca Pilar , Moreno Renata , Rojo Fernando
Published: July 2008

RNase R is a 3′-5′ highly processive exoribonuclease that can digest RNAs with extensive secondary structure. We analyzed the global effect of eliminating RNase R on thePseudomonas putidatranscriptome and the expression of thernrgene under diverse conditions. The absence of RNase R led to increased levels of many mRNAs, indicating that it plays an important role in mRNA turnover.
SoxAX Binding Protein, a Novel Component of the Thiosulfate-Oxidizing Multienzyme System in the Green Sulfur Bacterium
Chlorobium tepidum
Authors: Ogawa Takuro , Furusawa Toshinari , Nomura Ryohei , Seo Daisuke , Hosoya-Matsuda Naomi , Sakurai Hidehiro , Inoue Kazuhito
Published: July 2008

From the photosynthetic green sulfur bacterium
Chlorobium tepidum
(pro synon.
Chlorobaculum tepidum
), we have purified three factors indispensable for the thiosulfate-dependent reduction of the small, monoheme cytochrome
c
554
. These are homologues of sulfur-oxidizing (Sox) system factors found in various thiosulfate-oxidizing bacteria. The first factor is SoxYZ that serves as the acceptor for the reaction intermediates. The second factor is monomeric SoxB that is proposed to catalyze the hydrolytic cleavage of sulfate from the SoxYZ-bound oxidized product of thiosulfate. The third factor is the trimeric cytochrome
c
551
, composed of the monoheme cytochrome SoxA, the monoheme cytochrome SoxX, and the product of the hypothetical open reading frame CT1020. The last three components were expressed separately in
Escherichia coli
cells and purified to homogeneity. In the presence of the other two Sox factors, the recombinant SoxA and SoxX showed a low but discernible thiosulfate-dependent cytochrome
c
554
reduction activity. The further addition of the recombinant CT1020 protein greatly increased the activity, and the total activity was as high as that of the native SoxAX-CT1020 protein complex. The recombinant CT1020 protein participated in the formation of a tight complex with SoxA and SoxX and will be referred to as SAXB (SoxAX binding protein). Homologues of the SAXB gene are found in many strains, comprising roughly about one-third of the thiosulfate-oxidizing bacteria whose
sox
gene cluster sequences have been deposited so far and ranging over the
Chlorobiaciae, Chromatiaceae, Hydrogenophilaceae, Oceanospirillaceae
, etc. Each of the deduced SoxA and SoxX proteins of these bacteria constitute groups that are distinct from those found in bacteria that apparently lack SAXB gene homologues.
CDP-Alcohol Hydrolase, a Very Efficient Activity of the 5′-Nucleotidase/UDP-Sugar Hydrolase Encoded by the
ushA
Gene of
Yersinia intermedia
and
Escherichia coli
Authors: Alves-Pereira Isabel , Canales José , Cabezas Alicia , Martín Cordero Paloma , Costas María Jesús , Cameselle José Carlos
Published: July 2008

Nucleoside 5′-diphosphate-X hydrolases are interesting enzymes to study due to their varied activities and structure-function relationships and the roles they play in the disposal, assimilation, and modulation of the effects of their substrates. Few of these enzymes with a preference for CDP-alcohols are known. In
Yersinia intermedia
suspensions prepared from cultures on Columbia agar with 5% sheep blood, we found a CDP-alcohol hydrolase liberated to Triton X-100-containing medium. Growth at 25°C was deemed optimum in terms of the enzyme-activity yield. The purified enzyme also displayed 5′-nucleotidase, UDP-sugar hydrolase, and dinucleoside-polyphosphate hydrolase activities. It was identified as the protein product (UshA
Yi
) of the
Y. intermedia ushA
gene (
ushA
Yi
) by its peptide mass fingerprint and by PCR cloning and expression to yield active enzyme. All those activities, except CDP-alcohol hydrolase, have been shown to be the properties of UshA of
Escherichia coli
(UshA
Ec
). Therefore, UshA
Ec
was expressed from an appropriate plasmid and tested for CDP-alcohol hydrolase activity. UshA
Ec
and UshA
Yi
behaved similarly. Besides being the first study of a UshA enzyme in the genus
Yersinia
, this work adds CDP-alcohol hydrolase to the spectrum of UshA activities and offers a novel perspective on these proteins, which are viewed here for the first time as highly efficient enzymes with
k
cat
/K

m

ratios near the theoretical maximum level of catalytic activities. The results are discussed in the light of the known structures of UshA
Ec
conformers and the respective homology models constructed for UshA
Yi
, and also in relation to possible biological functions. Interestingly, every
Yersinia
species with a sequenced genome contains an intact
ushA
gene, except
Y. pestis
, which in all its sequenced biovars contains a
ushA
gene inactivated by frameshift mutations.
Translational Regulation by an Intramolecular Stem-Loop Is Required for Intermolecular RNA Regulation of theparAddiction Module
Authors: Shokeen Sonia , Patel Smita , Greenfield Tony J. , Brinkman Cassandra , Weaver Keith E.
Published: July 2008

Theparstability determinant ofEnterococcus faecalisplasmid pAD1 is the only antisense RNA-regulated addiction module identified to date in gram-positive bacteria.parencodes two small, convergently transcribed RNAs, designated RNA I and RNA II, that function as the toxin (Fst)-encoding and antitoxin components, respectively. Previous work showed that structures at the 5′ end of RNA I are important in regulating its translation. The work presented here reveals that a stem-loop sequestering the Fst ribosome binding site is required for translational repression but a helix sequestering the 5′ end of RNA I is not. Furthermore, disruption of the stem-loop prevented RNA II-mediated repression of Fst translation in vivo. Finally, although Fst-encoding wild-type RNA I is not toxic inEscherichia coli, mutations affecting stem-loop stability resulted in toxicity in this host, presumably due to increased translation.
Two Members of a Network of Putative Na
+
/H
+
Antiporters Are Involved in Salt and pH Tolerance of the Freshwater Cyanobacterium
Synechococcus elongatus
Authors: Billini Maria , Stamatakis Kostas , Sophianopoulou Vicky
Published: July 2008

Synechococcus elongatus
strain PCC 7942 is an alkaliphilic cyanobacterium that tolerates a relatively high salt concentration as a freshwater microorganism. Its genome sequence revealed seven genes,
nha1
to
nha7
(syn_pcc79420811, syn_pcc79421264, syn_pcc7942359, syn_pcc79420546, syn_pcc79420307, syn_pcc79422394, and syn_pcc79422186), and the deduced amino acid sequences encoded by these genes are similar to those of Na
+
/H
+
antiporters. The present work focused on molecular and functional characterization of these
nha
genes encoding Na
+
/H
+
antiporters. Our results show that of the
nha
genes expressed in
Escherichia coli
, only
nha3
complemented the deficient Na
+
/H
+
antiporter activity of the Na
+
-sensitive TO114 recipient strain. Moreover, two of the cyanobacterial strains with separate disruptions in the
nha
genes (Δ
nha1
, Δ
nha2
, Δ
nha3
, Δ
nha4
, Δ
nha5
, and Δ
nha7
) had a phenotype different from that of the wild type. In particular, Δ
nhA3
cells showed a high-salt- and alkaline-pH-sensitive phenotype, while Δ
nha2
cells showed low salt and alkaline pH sensitivity. Finally, the transcriptional profile of the
nha1
to
nha7
genes, monitored using the real-time PCR technique, revealed that the
nha6
gene is upregulated and the
nha1
gene is downregulated under certain environmental conditions.
Function of a Chemotaxis-Like Signal Transduction Pathway in Modulating Motility, Cell Clumping, and Cell Length in the Alphaproteobacterium
Azospirillum brasilense
Authors: Bible Amber N. , Stephens Bonnie B. , Ortega Davi R. , Xie Zhihong , Alexandre Gladys
Published: July 2008

A chemotaxis signal transduction pathway (hereafter called Che1) has been previously identified in the alphaproteobacterium
Azospirillum brasilens
e. Previous experiments have demonstrated that although mutants lacking CheB and/or CheR homologs from this pathway are defective in chemotaxis, a mutant in which the entire chemotaxis pathway has been mutated displayed a chemotaxis phenotype mostly similar to that of the parent strain, suggesting that the primary function of this Che1 pathway is not the control of motility behavior. Here, we report that mutants carrying defined mutations in the
cheA1
(strain AB101) and the
cheY1
(strain AB102) genes and a newly constructed mutant lacking the entire operon [Δ(
cheA1-cheR1
)::Cm] (strain AB103) were defective, but not null, for chemotaxis and aerotaxis and had a minor defect in swimming pattern. We found that mutations in genes of the Che1 pathway affected the cell length of actively growing cells but not their growth rate. Cells of a mutant lacking functional
cheB1
and
cheR1
genes (strain BS104) were significantly longer than wild-type cells, whereas cells of mutants impaired in the
cheA1
or
cheY1
genes, as well as a mutant lacking a functional Che1 pathway, were significantly shorter than wild-type cells. Both the modest chemotaxis defects and the observed differences in cell length could be complemented by expressing the wild-type genes from a plasmid. In addition, under conditions of high aeration, cells of mutants lacking functional
cheA1
or
cheY1
genes or the Che1 operon formed clumps due to cell-to-cell aggregation, whereas the mutant lacking functional CheB1 and CheR1 (BS104) clumped poorly, if at all. Further analysis suggested that the nature of the exopolysaccharide produced, rather than the amount, may be involved in this behavior. Interestingly, mutants that displayed clumping behavior (lacking
cheA1
or
cheY1
genes or the Che1 operon) also flocculated earlier and quantitatively more than the wild-type cells, whereas the mutant lacking both CheB1 and CheR1 was delayed in flocculation. We propose that the Che1 chemotaxis-like pathway modulates the cell length as well as clumping behavior, suggesting a link between these two processes. Our data are consistent with a model in which the function of the Che1 pathway in regulating these cellular functions directly affects flocculation, a cellular differentiation process initiated under conditions of nutritional imbalance.
The Metal Dependence of Pyoverdine Interactions with Its Outer Membrane Receptor FpvA
Authors: Greenwald Jason , Zeder-Lutz Gabrielle , Hagege Agnès , Celia Hervé , Pattus Franc
Published: July 2008

To acquire iron,Pseudomonas aeruginosasecretes the fluorescent siderophore pyoverdine (Pvd), which chelates iron and shuttles it into the cells via the specific outer membrane transporter FpvA. We studied the role of iron and other metals in the binding and transport of Pvd by FpvA and conclude that there is no significant affinity between FpvA and metal-free Pvd. We found that the fluorescent in vivo complex of iron-free FpvA-Pvd is in fact a complex with aluminum (FpvA-Pvd-Al) formed from trace aluminum in the growth medium. WhenPseudomonas aeruginosawas cultured in a medium that had been treated with a metal affinity resin, the in vivo formation of the FpvA-Pvd complex and the recycling of Pvd on FpvA were nearly abolished. The accumulation of Pvd in the periplasm ofPseudomonas aeruginosawas also reduced in the treated growth medium, while the addition of 1 μM AlCl3to the treated medium restored the effects of trace metals observed in standard growth medium. Using fluorescent resonance energy transfer and surface plasmon resonance techniques, the in vitro interactions between Pvd and detergent-solubilized FpvA were also shown to be metal dependent. We demonstrated that FpvA binds Pvd-Fe but not Pvd and that Pvd did not compete with Pvd-Fe for FpvA binding. In light of our finding that the Pvd-Al complex is transported across the outer membrane ofPseudomonas aeruginosa, a model for siderophore recognition based on a metal-induced conformation followed by redox selectivity for iron is discussed.
The
Bacillus cereus
GerN and GerT Protein Homologs Have Distinct Roles in Spore Germination and Outgrowth, Respectively
Authors: Senior Adam , Moir Anne
Published: July 2008

The GerT protein of
Bacillus cereus
shares 74% amino acid identity with its homolog GerN. The latter is a Na
+
/H
+
-K
+
antiporter that is required for normal spore germination in inosine. The germination properties of single and double mutants of
B. cereus
ATCC 10876 reveal that unlike GerN, which is required for all germination responses that involve the GerI germinant receptor, the GerT protein does not have a significant role in germination, although it is required for the residual GerI-mediated inosine germination response of a
gerN
mutant. In contrast, GerT has a significant role in outgrowth;
gerT
mutant spores do not outgrow efficiently under alkaline conditions and outgrow more slowly than the wild type in the presence of high NaCl concentrations. The GerT protein in
B. cereus
therefore contributes to the success of spore outgrowth from the germinated state during alkaline or Na
+
stress.
Characterization of Sec-Translocon-Dependent Extracytoplasmic Proteins of
Rickettsia typhi
Authors: Ammerman Nicole C. , Rahman M. Sayeedur , Azad Abdu F.
Published: July 2008

As obligate intracellular, vector-borne bacteria, rickettsiae must adapt to both mammalian and arthropod host cell environments. Deciphering the molecular mechanisms of the interactions between rickettsiae and their host cells has largely been hindered by the genetic intractability of these organisms; however, research in other gram-negative pathogens has demonstrated that many bacterial determinants of attachment, entry, and pathogenesis are extracytoplasmic proteins. The annotations of several rickettsial genomes indicate the presence of homologs of the Sec translocon, the major route for bacterial protein secretion from the cytoplasm. For
Rickettsia typhi
, the etiologic agent of murine typhus, homologs of the Sec-translocon-associated proteins LepB, SecA, and LspA have been functionally characterized; therefore, the
R. typhi
Sec apparatus represents a mechanism for the secretion of rickettsial proteins, including virulence factors, into the extracytoplasmic environment. Our objective was to characterize such Sec-dependent
R. typhi
proteins in the context of a mammalian host cell infection. By using the web-based programs LipoP, SignalP, and Phobius, a total of 191
R. typhi
proteins were predicted to contain signal peptides targeting them to the Sec translocon. Of these putative signal peptides, 102 were tested in an
Escherichia coli
-based alkaline phosphatase (PhoA) gene fusion system. Eighty-four of these candidates exhibited signal peptide activity in
E. coli
, and transcriptional analysis indicated that at least 54 of the
R. typhi
extracytoplasmic proteins undergo active gene expression during infections of HeLa cells. This work highlights a number of interesting proteins possibly involved in rickettsial growth and virulence in mammalian cells.
The Type III Pantothenate Kinase Encoded by
coaX
Is Essential for Growth of
Bacillus anthracis
Authors: Paige Carleitta , Reid Sean D. , Hanna Philip C. , Claiborne Al
Published: July 2008

In
Bacillus anthracis
, the novel type III pantothenate kinase (PanK

Ba

; encoded by
coaX
) catalyzes the first committed step in coenzyme A biosynthesis. We have demonstrated by analyzing the growth characteristics of a conditional
coaX
mutant that PanK

Ba

is an essential enzyme, thus contributing to its validation as a new antimicrobial target.
Doctors with difficulties: why so few women?
Authors: Firth-Cozens J
Published: July 2008

The National Clinical Assessment Service (NCAS), an NHS organisation that assesses doctors and dentists referred to them because of perceived difficulties, has produced a report describing data arising from its first 4 years, showing that male doctors were referred to the service considerably more often than female doctors. Despite women accounting for 42% of the general practitioner medical workforce and 37% of the medical hospital and community (H&C) workforce in 2004, only 13% of GPs and 20% of H&C NCAS referrals were women. When the H&C data were split into specialties, women were under-represented proportionally in all specialties. This paper offers a review of possible reasons for these gender differences and in doing so contributes to the debates concerning problems in performance and also the costs of employing a growing proportion of women doctors. Firstly, it hypothesises that the NCAS data may be non-representative of similar agency data, but finds that in disciplinary organisations of various types around the world, men are consistently over-represented. Secondly, it suggests that perhaps men are referred to such agencies more often than women because their employers are more lenient on women. There is no evidence for this, and it requires primary research to investigate it further. Finally, it considers gender differences in the attributes, beyond technical skills, that underpin a good doctor–patient relationship and finds that, on these attributes, women usually excel over men. In addition, far fewer women are disciplined for addiction. The implications of this for education and rehabilitation are considered. It concludes that any analysis of the economic costs of employing a greater proportion of female doctors must take into account the higher costs of men's litigation, discipline and retraining.
Concentric and radial joint systems within basic sills and their associated porosity enhancement, Neuquén Basin, Argentina
Authors: Bermúdez Adriana , Delpino Daniel H.
Published: July 2008

One of the remarkable characteristics of the Neuquén Basin is that a group of Cenozoic shallow microgabbroic sills exhibit gas and oil shows, and some have produced hydrocarbons for up to 20 years. Their capacity to act as reservoirs is caused by the increase in the effective porosity owing to the simultaneous development of petrological and fracture processes during cooling that generate systematic variations inside the intrusive bodies.

In this work we describe in detail an unusual intracolumnar joint system formed by two joint sets, referred to here as concentric and radial joint sets. This atypical joint system is developed within the small stacked polyhedrons into which certain zones of classic prismatic columns are transformed during the cooling processes. These small polyhedrons are laterally bounded by the vertical planes that limit the columns, and at the top and bottom by transversal joint planes oriented almost perpendicular to the column axis.

As a result of detailed macroscopic analysis, the genesis of the concentric joint sets have been interpreted as resulting from either the development of natural convective flows or from complex processes of cooling influenced by the joint-bounded planes of the columns. When the radial joint set is present, it is clearly related to the development of thermal stresses between the outer and inner parts of the small polyhedrons.

In geology textbooks and previous published papers the genesis of what is informally called ‘onion skin’ joint sets in basic intrusive and extrusive igneous rocks has been attributed to processes related to either weathering or deuteric alteration. However, based on detailed macroscopic observations, our work suggests that, in most cases, any type of alteration process only accentuates the planes previously generated during the cooling process.
Lithological and structural controls on the emplacement and morphology of sills in sedimentary basins
Authors: Thomson Ken , Schofield Nick
Published: July 2008

Three-dimensional seismic data from the NW European Atlantic margin provide detailed data on the relationships between sills, dykes, laccoliths and pre-existing basin structure. The data show that sills are predominantly concave-upwards in shape, being complete or partial versions of radially or bilaterally symmetrical forms that possess flat inner saucers connected to a flat outer rim by a steeply inclined sheet. Sills can be sourced from the steep climbing portions of deeper sills, and can be shown to exploit pre-existing faults to feed shallower sills and laccoliths. Magma flow patterns, as revealed by opacity rendering, suggest that sills propagate upwards and outwards away from the magma feeder. As an individual sill can consist of several leaves emplaced at different stratigraphic levels, and as a sill or dyke can provide magma to other sills and laccoliths, the data suggest that neutral buoyancy concepts may not provide a complete explanation for the mechanism and level of sill emplacement. Instead, the data suggest that the presence of ductile horizons such as overpressured shales may permit sill formation below the neutrally buoyant level. Emplacement below the level of neutral buoyancy would permit sill inflation, and the associated fracturing of the country rock would permit magma to exploit these pathways in order to ascend to shallower levels and feed shallower intrusions.