bluuebunny/crossref_metadata_2025 huggingface dataset:
Determinants of Complementary Medicine Service Utilization in a Regional South Australian Population: A Cross‐Sectional Study
Authors: Leach Matthew J
Published: March 2020

Purpose

The dearth of accessible and appropriate conventional health care services in regional areas may drive many people to seek care provided by complementary medicine (CM) practitioners. Understanding the drivers of CM service use in this population may help inform future health services planning. This study set out to examine the determinants of CM service utilization in a regional South Australian population.

Methods

Adults residing in regional South Australia between April 2017 and March 2018 were invited to complete the 44‐item consumer utilization, expectations, and experiences of health care instrument. The questionnaire was available in print and online, and was promoted using a multimodal recruitment campaign. Independent predictors of CM service use were determined using multivariate logistic regression.

Findings

Completing the questionnaire were 3,926 adults (56.7% aged 50+ years; 52.5% females). Forty‐seven percent of participants reported using a CM service within the last 12 months. Univariate analyses revealed a statistically significant association between 13 factors and CM service use. When these factors were forced into a multivariate logistic regression model, the number of predictors of CM service utilization was reduced to 10, which included 6 predisposing factors, 3 enabling factors, and 1 personal health practice. This model was able to explain 23.1% (Nagelkerke R2 = 0.231) of the variation in CM service use in this sample.

Conclusions

A high level of CM service use was reported among participants living in regional South Australia. The findings highlight the degree to which the appropriateness of health services impacts health‐seeking behavior in regional communities.
Myristate and the ecology of AM fungi: significance, opportunities, applications and challenges
Authors: Rillig Matthias C. , Aguilar‐Trigueros Carlos A. , Anderson Ian C. , Antonovics Janis , Ballhausen Max‐Bernhard , Bergmann Joana , Bielcik Milos , Chaudhary V. Bala , Deveautour Coline , Grünfeld Leonie , Hempel Stefan , Lakovic Milica , Lammel Daniel R. , Lehmann Anika , Lehmann Johannes , Leifheit Eva F. , Liang Yun , Li Erqin , Lozano Yudi M. , Manntschke Annette , Mansour India , Oviatt Peter , Pinek Liliana , Powell Jeff R. , Roy Julien , Ryo Masahiro , Sosa‐Hernández Moisés A. , Veresoglou Stavros D. , Wang Dongwei , Yang Gaowen , Zhang Haiyang
Published: March 2020

A recent study by Sugiura and coworkers reported the non‐symbiotic growth and spore production of an arbuscular mycorrhizal (AM) fungus, Rhizophagus irregularis, when the fungus received an external supply of certain fatty acids, myristates (C:14). This discovery follows the insight that AM fungi receive fatty acids from their hosts when in symbiosis. If this result holds up and can be repeated under nonsterile conditions and with a broader range of fungi, it has numerous consequences for our understanding of AM fungal ecology, from the level of the fungus, at the plant community level, and to functional consequences in ecosystems. In addition, myristate may open up several avenues from a more applied perspective, including improved fungal culture and supplementation of AM fungi or inoculum in the field. We here map these potential opportunities, and additionally offer thoughts on potential risks of this potentially new technology. Lastly, we discuss the specific research challenges that need to be overcome to come to an understanding of the potential role of myristate in AM ecology.
The pulse of the tree is under genetic control: eucalyptus as a case study
Authors: Bartholomé Jérôme , Mabiala André , Burlett Régis , Bert Didier , Leplé Jean‐Charles , Plomion Christophe , Gion Jean‐Marc
Published: March 2020

The pulse of the tree (diurnal cycle of stem radius fluctuations) has been widely studied as a way of analyzing tree responses to the environment, including the phenotypic plasticity of tree–water relationships in particular. However, the genetic basis of this daily phenotype and its interplay with the environment remain largely unexplored. We characterized the genetic and environmental determinants of this response, by monitoring daily stem radius fluctuation (dSRF) on 210 trees from a Eucalyptus urophylla × E. grandis full‐sib family over 2 years. The dSRF signal was broken down into hydraulic capacitance, assessed as the daily amplitude of shrinkage (DA), and net growth, estimated as the change in maximum radius between two consecutive days (ΔR). The environmental determinants of these two traits were clearly different: DA was positively correlated with atmospheric variables relating to water demand, while ΔR was associated with soil water content. The heritability for these two traits ranged from low to moderate over time, revealing a time‐dependent or environment‐dependent complex genetic determinism. We identified 686 and 384 daily quantitative trait loci (QTL) representing 32 and 31 QTL regions for DA and ΔR, respectively. The identification of gene networks underlying the 27 major genomics regions for both traits generated additional hypotheses concerning the biological mechanisms involved in response to water demand and supply. This study highlights that environmentally induced changes in daily stem radius fluctuation are genetically controlled in trees and suggests that these daily responses integrated over time shape the genetic architecture of mature traits.
Reviewer Acknowledgements
Authors: Willianms Ellery
Published: March 2020

Business and Management Studies (BMS) would like to acknowledge the following reviewers for their assistance with peer review of manuscripts for this issue. Many authors, regardless of whether BMS publishes their work, appreciate the helpful feedback provided by the reviewers. Their comments and suggestions were of great help to the authors in improving the quality of their papers. Each of the reviewers listed below returned at least one review for this issue. Reviewers for Volume 6, Number 1 Ashford Chea, Benedict College, USABahram Abediniangerabi, University of Texas, USADalia Susniene, Kaunas University of Technology, LithuaniaFlorin Peci, University of Peja, KosovoGabriela O. Chiciudean, University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca, RomaniaHung-Che Wu, Nanfang College of Sun Yat-sen University, ChinaJulia Stefanova, Bulgarian Academy of Sciences, BulgariaMichael Okoche, University of South Africa, UgandaMike Rayner, University of Portsmouth, UKSandeep Kumar, Tecnia Institute of Advanced Studies, Affiliated to GGSIP University Delgi, IndiaTetiana Paientko, Kyiv National Economic Univercity, UkraineZoran Mastilo, University of East Sarajevo, Bosnia and Herzegovina Ellery WillianmsEditorial AssistantOn behalf of,The Editorial Board of Business and Management StudiesRedfame Publishing9450 SW Gemini Dr. #99416Beaverton, OR 97008, USAURL: http://bms.redfame.com
Foreign Language Competences of Educators and Areas of Their Application
Authors: Adamczyk Roman
Published: March 2020

The present article provides an overview of key areas of educators’ language skills with an emphasis on foreign-language competence. Foreign language competences of educators at all levels of schooling constitute an integral part of the contemporary pedagogical activity, being historically embedded in both Czech and foreign linguistic and language teaching cultures. The latter competences form a subset of general and ´soft´ skills in the area of communication and apowerful tool of knowledge acquisition, enabling access to foreign language resources in libraries, databases and the whole of the internet platform, as well as communication in interactive social settings of conferences and other professional contexts. The article primarily focuses on foreign language competences of non-language teachers as a part of their professional erudition and on the areas of application of both productive and receptive skills, including speaking, writing, listening, reading and translation. The convergence of language teaching and content subjects resulted in the emergence of new didactic approaches that further underscore the importance and potential of foreign language competence in the professional activity of teachers.
Measurement of the Natural Convection Heat Transfer in a Magnesium Oxide Electrolytic Cell Concept
Authors: Venstrom Luke J. , Yager Jacob , Vervynckt Todd , Ogland-Hand Jonathan D. , Nudehi Shahin S.
Published: March 2020

The rate of heat transfer by natural convection between the wall and electrolyte of an electrolytic cell that produces magnesium (Mg) from magnesium oxide (MgO) at temperatures near 1000 °C in a molten fluoride salt electrolyte is presented. An experimental model of the cell was developed that enabled measurements of the heat transfer in the absence of electrolysis and at temperatures less than 100 °C over ranges of Rayleigh numbers from 1 × 10−7 to 7 × 10−8 and Prandtl numbers from 2 to 6200, ranges that include those anticipated in the operation of the MgO electrolytic cell. The model avoids the substantial experimental challenges associated with the high-temperature, corrosive molten salt to enable a conservative estimate of the heat transfer at a lower cost and greater accuracy than would otherwise be possible. The results are correlated by the expression Nu = 0.412Ra0.23Pr0.15 with Nusselt numbers spanning 30–80. The application of the correlation shows that the heat transfer between the cell wall and the molten fluoride electrolyte at ≈1000 °C is characterized by convection coefficients between 100 and 600 W/m2-K and is fast enough to enable heat fluxes up to 10 W/cm2 without compromising the structural integrity of the steel cell wall.
Heating Analysis of a Water Droplet in Between Multi-Wall Hydrophobic Surfaces
Authors: Al-Sharafi Abdullah , Yilbas Bekir S. , Sahin Ahmet Z. , Al-Qahtani Hussain
Published: March 2020

Droplet heat transfer in between parallelly located superhydrophobic plates is examined. The thermal field inside the droplet is predicted by adopting the experimental conditions. The influence of plates spacing (heights) on the thermal response of the droplet fluid is investigated. Particle injection velocimetry (PIV) is used to validate the velocity predictions. We demonstrated that predictions of flow velocity are in agreement with those of the PIV results. The heating of the droplet in the absence of the top plate gives four circulation cells in the droplet. Once the top superhydrophobic plate is introduced, the flow structure alters, and the number of the circulating structures reduces to two. Lowering the height of the plates increases the droplet Laplace pressure while modifying the fluid flow and thermal behavior. The Bond number is lower than one for all the cases considered; hence, demonstrating that the Marangoni force affects the formation of the circulation cells. The cells redistribute the heated fluid in the droplet interior, which is clearly apparent for the plates with small heights. Temperature enhancement in the droplet bottom section is attributed to the flow current formed due to heat diffusion. The Nusselt number corresponding to the bottom plate increases as the plate heights reduces; however, the opposite is true for that corresponding to the top plate.
Effect of low frequency power on the electron energy distribution function in argon inductively coupled plasmas
Authors: Kim Ju-Ho , Kwon Deuk-Chul , Chung Chin-Wook
Published: March 2020

In plasma processing and application, the electron energy distribution function (EEDF) is of fundamental interest because the ion and radical densities related to physical and chemical reactions on the substrate are predominantly governed by the EEDF or electron temperature. In this paper, the effect of low frequency power on the EEDF is investigated when 2 MHz power is added to the plasma originally driven at 13.56 MHz. In a 13.56 MHz operation, the EEDF shows a Maxwellian-like distribution, and as the RF power increases, the electron density increases and the electron temperature decreases. However, when a small amount of 2 MHz power is applied to the 13.56 MHz discharge, the electron density slightly increases and the electron temperature significantly increases. In dual-frequency operation, EEDFs have a low slope of low-energy region and evolve into a Druyvesteyn-like distribution. It turns out that the dual-frequency operation can significantly change the electron temperature. This is consistent with the results calculated using the analytical electron heating model, and the relevant heating mechanism is also presented.
Medium amplitude parallel superposition (MAPS) rheology. Part 1: Mathematical framework and theoretical examples
Authors: Lennon Kyle R. , McKinley Gareth H. , Swan James W.
Published: March 2020

A new mathematical representation for nonlinear viscoelasticity is presented based on the application of the Volterra series expansion to the general nonlinear relationship between shear stress and shear strain history. This theoretical and experimental framework, which we call medium amplitude parallel superposition (MAPS) rheology, reveals a new material property, the third-order complex modulus, which describes completely the weakly nonlinear response of a viscoelastic material in an arbitrary simple shear flow. In this first part, we discuss several theoretical aspects of this mathematical formulation and new material property. For example, we show how MAPS measurements can be performed in strain- or stress-controlled contexts and provide relationships between the weakly nonlinear response functions measured in each case. We show that the MAPS response function is a superset of the response functions that have been previously reported in medium amplitude oscillatory shear and parallel superposition rheology experiments. We also show how to exploit inherent symmetries of the MAPS response function to reduce it to a minimal domain for straightforward measurement and visualization. We compute this material property for a few constitutive models to illustrate the potential richness of the datasets generated by MAPS experiments. Finally, we discuss the MAPS framework in the context of some other nonlinear, time-dependent rheological probes and explain how the MAPS methodology has a distinct advantage over these others because it generates data embedded in a very high-dimensional space without driving fluid mechanical instabilities, and is agnostic to the flow protocol.
Decreased Resistance to Primary IntravenousCryptococcus neoformansInfection in Aged Mice despite Adequate Resistance to Intravenous Rechallenge
Authors: Aguirre Karen M. , Gibson George W. , Johnson Lawrence L.
Published: March 2020

It is often stated that impaired immune functions in the aged underlie their greater susceptibility to infections. Indeed, in many experimental settings, T-cell responses in aged mice have been shown to be deficient compared with those from young adults. Nonetheless, there are very few examples where a greater susceptibility to infection in aged mice has been demonstrated to result from impaired T-cell function. The clinical importance of understanding the basis for increased susceptibility to infection that accompanies advanced age dictates a need for experimental models with which to study the effect that aging has on immunological resistance to infection. This study was undertaken to investigate whether aged mice were less resistant than young adult control mice to infection with the fungusCryptococcus neoformans. After a primary intravenous challenge with yeast, aged mice died sooner and developed higher organ burdens of yeast than did young adults. Deficient in vitro responses were observed in T cells from aged mice; however, greater susceptibility to intravenous infection appeared not to result from less effective T-cell-dependent resistance in vivo. In fact, T-cell-replete aged mice were more susceptible to intravenous cryptococcal infection than were T-cell-depleted young adults. Furthermore, aged mice were as resistant to primary pulmonary challenge withCryptococcusas were young adults. Similarly, vaccinated aged mice were as resistant to rechallenge as were young adult counterparts. Therefore, despite demonstrably deficient in vitro responses of T cells from aged mice, their T-cell-dependent resistance toC. neoformansis as effective as that of young adults.
Characterization of an Acidic-pH-Inducible Stress Protein (hsp70), a Putative Sulfatide Binding Adhesin, fromHelicobacter pylori
Authors: Huesca Mario , Goodwin Avery , Bhagwansingh Arianna , Hoffman Paul , Lingwood Clifford A.
Published: March 2020

The in vitro glycolipid binding specificity of the gastric pathogenHelicobacter pyloriis altered to include sulfated glycolipids (sulfatides) following brief exposure of the organism to acid pH typical of the stomach. This change is prevented by anti-hsp70 antibodies, suggesting that hsp70 may be a stress-induced surface adhesin, mediating sulfatide recognition. To facilitate investigation of the role of hsp70 in attachment, we have cloned and sequenced theH. pylori hsp70gene (dnaK). Thehsp70gene was identified by probing a cosmid DNA library made fromH. pylori439 with a PCR amplicon generated with oligonucleotides synthesized to highly conserved regions ofdnaK. The 1.9-kbH. pylori hsp70gene encodes a product of 616 amino acids. Primer extension analysis revealed a single transcription start site, while Northern blot analysis established thathsp70was preferentially induced by low pH rather than by heat shock. The ability ofH. pylorito alter its glycolipid binding specificity following exposure to low pH by upregulatinghsp70and by expressing hsp70 on the bacterial surface may provide a survival advantage during periods of high acid stress.
The Dominant Epitope ofBorrelia gariniiOuter Surface Protein C Recognized by Sera from Patients with Neuroborreliosis Has a Surface-Exposed Conserved Structural Motif
Authors: Mathiesen Marianne J. , Holm Arne , Christiansen Michael , Blom Jens , Hansen Klaus , Østergaard Søren , Theisen Michael
Published: March 2020

Epitope mapping of outer surface protein C (OspC) by using sera from patients with neuroborreliosis led to the identification of one single major immunodominant epitope within the C-terminal 10 amino acid residues. Peptide binding studies and alanine replacement scanning of the C-terminal decapeptide, PVVAESPKKP, revealed a critical role for the PKKP sequence and its terminal carboxyl group for the binding of immunoglobulin M (IgM) antibodies from patients with Lyme borreliosis. Electron microscopy of antibody-labeled spirochetes indicated that the C-terminal region is exposed on the surface of the spirochete. Based on homology to proteins of known function, this region most probably adopts a polyproline II-like helix, which is found in surface-exposed structures involved in protein-protein interactions. This structural motif is highly conserved inBorreliaspecies causing Lyme borreliosis and subjected to purifying selection. We suggest that the abundance of the C-terminal region of OspC on the surface ofB. burgdorferiallows a multimeric high-avidity interaction between the spirochete and surface Igs on B cells. The resulting cross-linking of surface Igs on B cells may induce a T-cell-independent B-cell activation without IgM-to-IgG switching, thus explaining the lack of IgG antibodies to OspC in neuroborreliosis.
Low Endotoxic Potential ofLegionella pneumophilaLipopolysaccharide due to Failure of Interaction with the Monocyte Lipopolysaccharide Receptor CD14
Authors: Neumeister B. , Faigle M. , Sommer M. , Zähringer U. , Stelter F. , Menzel R. , Schütt C. , Northoff H.
Published: March 2020

Legionella pneumophila, a gram-negative bacterium causing Legionnaires’ disease and Pontiac fever, was shown to be highly reactive in in vitro gelation ofLimuluslysate but not able to induce fever and the local Shwartzman reaction in rabbits and mice. We analyzed the capacity of purifiedL. pneumophilalipopolysaccharide (LPS-Lp) to induce activation of the human monocytic cell line Mono Mac 6, as revealed by secretion of proinflammatory cytokines and desensitization to subsequent LPS stimulation. We showed that despite normal reactivity of LPS-Lp in theLimulusamoebocyte lysate assay, induction of cytokine secretion in Mono Mac 6 cells and desensitization to an endotoxin challenge required LPS-Lp concentrations 1,000 times higher than for LPS ofSalmonella entericaserovar Minnesota. Therefore, we examined the interaction of LPS-Lp with the LPS receptor CD14. We demonstrated that LPS-Lp did not bind to membrane-bound CD14 expressed on transfected CHO cells, nor did it react with soluble CD14. Our results suggest that the low endotoxic potential of LPS-Lp is due to a failure of interaction with the LPS receptor CD14.
Ferrous Iron Uptake in
Cryptococcus neoformans
Authors: Jacobson Eric S. , Goodner Asha Prasad , Nyhus Karin J.
Published: March 2020

Previous studies have implicated ferric reduction in the iron uptake pathway of the opportunistic pathogen
Cryptococcus neoformans
. Here we studied iron uptake directly, using
55
Fe in the presence of reductants. Uptake was linear with respect to time and number of yeast cells. The plot of uptake versus concentration exhibited a steep rise up to about 1 μM, a plateau between 1 and 25 μM, and a second steep rise above 25 μM, consistent with high- and low-affinity uptake systems. A

K
m

for high-affinity uptake was estimated to be 0.6 μM Fe(II); 1 μM was used for standardized uptake assays. At this concentration, the uptake rate was 110 ± 3 pmol/10
6
cells/h. Iron repletion (15 μM) and copper starvation drastically decreased high-affinity iron uptake. Incubation at 0°C or in the presence of 2 mM KCN abolished high-affinity iron uptake, suggesting that uptake requires metabolic energy. When exogenous reducing agents were not supplied and the culture was washed free of secreted reductants, uptake was reduced by 46%; the remaining uptake activity presumably was dependent upon the cell membrane ferric reductase. Further decreases in free Fe(II) levels achieved by trapping with bathophenanthroline disulfonate or reoxidizing with potassium nitrosodisulfonate reduced iron uptake very drastically, suggesting that it is the Fe(II) species which is transported by the high-affinity transporter. The uptake of Fe was stimulated two- to threefold by deferoxamine, but this increment could be abolished by copper starvation or inhibition of the ferric reductase by Pt, indicating that Fe solubilized by this molecule also entered the reductive iron uptake pathway.
Targeted Disruption of theGRA2Locus inToxoplasma gondiiDecreases Acute Virulence in Mice
Authors: Mercier Corinne , Howe Daniel K. , Mordue Dana , Lingnau Maren , Sibley L. David
Published: March 2020

Following invasion into the host cell, the protozoanToxoplasma gondiisecretes a variety of proteins that modify the parasitophorous vacuole. Within the vacuole, the 28-kDa dense granule protein known as GRA2 is specifically targeted to the tubulovesicular network which forms connections with the vacuolar membrane. To investigate the importance of GRA2, we derived from strain RH a mutantT. gondiiline in whichGRA2was disrupted by replacement with the marker Ble (selecting for phleomycin resistance). The Δgra2mutant invaded and grew normally in both fibroblasts and macrophages in vitro; however, it was less virulent during acute infection in mice. The survival rate of mice inoculated with Δgra2was significantly higher; some infected mice survived the acute infection, whereas all mice infected with the wild-type strain RH succumbed to early death. Chronic infection by Δgra2was detected by positive serology, immunohistochemical detection of parasites and cysts in the brain, and reisolation of parasites by bioassay at 6 weeks postinfection. Thus, absence of GRA2 partially attenuates the virulence ofT. gondiiduring the acute phase of infection and allows for establishment of chronic infection by the otherwise highly virulent RH strain. These results establish that GRA2 plays an important role during in vivo infection and provide a potential model for examining acute pathogenesis byT. gondii.
Molecular Cloning, Expression, and Immunogenicity of MTB12, a Novel Low-Molecular-Weight Antigen Secreted by
Mycobacterium tuberculosis
Authors: Webb John R. , Vedvick Thomas S. , Alderson Mark R. , Guderian Jeffrey A. , Jen Shyian S. , Ovendale Pamela J. , Johnson Stephen M. , Reed Steven G. , Skeiky Yasir A. W.
Published: March 2020

Proteins secreted into the culture medium by
Mycobacterium tuberculosis
are thought to play an important role in the development of protective immune responses. In this report, we describe the molecular cloning of a novel, low-molecular-weight antigen (MTB12) secreted by
M. tuberculosis
. Sequence analysis of the MTB12 gene indicates that the protein is initially synthesized as a 16.6-kDa precursor protein containing a 48-amino-acid hydrophobic leader sequence. The mature, fully processed form of MTB12 protein found in culture filtrates has a molecular mass of 12.5 kDa. MTB12 protein constitutes a major component of the
M. tuberculosis
culture supernatant and appears to be at least as abundant as several other well-characterized culture filtrate proteins, including members of the 85B complex. MTB12 is encoded by a single-copy gene which is present in both virulent and avirulent strains of the
M. tuberculosis
complex, the BCG strain of
M. bovis
, and
M. leprae
. Recombinant MTB12 containing an N-terminal six-histidine tag was expressed in
Escherichia coli
and purified by affinity chromatography. Recombinant MTB12 protein elicited in vitro proliferative responses from the peripheral blood mononuclear cells of a number of purified protein derivative-positive (PPD
+
) human donors but not from PPD
−
donors.
Characterization of Major Surface Glycoprotein Genes of HumanPneumocystis cariniiand High-Level Expression of a Conserved Region
Authors: Mei Qin , Turner Ross E. , Sorial Vivian , Klivington Diane , Angus C. William , Kovacs Joseph A.
Published: March 2020

To facilitate studies ofPneumocystis cariniiinfection in humans, we undertook to better characterize and to express the major surface glycoprotein (MSG) of humanP. carinii, an important protein in host-pathogen interactions. Seven MSG genes were cloned from a single isolate by PCR or genomic library screening and were sequenced. The predicted proteins, like rat MSGs, were closely related but unique variants, with a high level of conservation among cysteine residues. A conserved immunodominant region (of approximately 100 amino acids) near the carboxy terminus was expressed at high levels inEscherichia coliand used in Western blot studies. All 49 of the serum samples, which were taken from healthy controls as well as from patients with and withoutP. cariniipneumonia, were reactive with this peptide by Western blotting, supporting the hypothesis that most adult humans have been infected withP. cariniiat some point. This recombinant MSG fragment, which is the first humanP. cariniiantigen available in large quantities, may be a useful reagent for investigating the epidemiology ofP. cariniiinfection in humans.
Comparison of Sample Sequences of the
Salmonella typhi
Genome to the Sequence of the Complete
Escherichia coli
K-12 Genome
Authors: McClelland Michael , Wilson Richard K.
Published: March 2020

Raw sequence data representing the majority of a bacterial genome can be obtained at a tiny fraction of the cost of a completed sequence. To demonstrate the utility of such a resource, 870 single-stranded M13 clones were sequenced from a shotgun library of the
Salmonella typhi
Ty2 genome. The sequence reads averaged over 400 bases and sampled the genome with an average spacing of once every 5,000 bases. A total of 339,243 bases of unique sequence was generated (approximately 7% representation). The sample of 870 sequences was compared to the complete
Escherichia coli
K-12 genome and to the rest of the GenBank database, which can also be considered a collection of sampled sequences. Despite the incomplete
S. typhi
data set, interesting categories could easily be discerned. Sixteen percent of the sequences determined from
S. typhi
had close homologs among known
Salmonella
sequences (
P
< 1
e
−40
in BlastX or BlastN), reflecting the proportion of these genomes that have been sequenced previously; 277 sequences (32%) had no apparent orthologs in the complete
E. coli
K-12 genome (
P
> 1
e
−20
), of which 155 sequences (18%) had no close similarities to any sequence in the database (
P
> 1
e
−5
). Eight of the 277 sequences had similarities to genes in other strains of
E. coli
or plasmids, and six sequences showed evidence of novel phage lysogens or sequence remnants of phage integrations, including a member of the lambda family (
P
< 1
e
−15
). Twenty-three sample sequences had a significantly closer similarity a sequence in the database from organisms other than the
E. coli/Salmonella
clade (which includes
Shigella
and
Citrobacter
). These sequences are new candidate lateral transfer events to the
S. typhi
lineage or deletions on the
E. coli
K-12 lineage. Eleven putative junctions of insertion/deletion events greater than 100 bp were observed in the sample, indicating that well over 150 such events may distinguish
S. typhi
from
E. coli
K-12. The need for automatic methods to more effectively exploit sample sequences is discussed.
Partially Protective Vaccination Permits the Development of Latency in a Normally Virulent Strain ofToxoplasma gondii
Authors: Yap George S. , Scharton-Kersten Tanya , Ferguson David J. P. , Howe Dan , Suzuki Yasuhiro , Sher Alan
Published: March 2020

The virulent RH strain ofToxoplasma gondiiis acutely lethal in mice and fails to establish chronic infection. Vaccination of BALB/c mice with a soluble tachyzoite antigen preparation, STAg, in combination with the immunostimulatory cytokine interleukin-12 results in partial protection against RH lethal challenge. Nevertheless, brain tissue obtained from surviving, vaccinated mice as late as 1 year after RH infection contained latent parasite forms as demonstrated by subinoculation into naive recipients. The tachyzoites arising in the subinoculated animals were genetically indistinguishable from the original RH inoculum. Microscopic examination revealed that the persistent parasite forms present in the brains of vaccinated and challenged mice have a tissue cyst-like morphology and express the bradyzoite antigen BAG-1 but not the tachyzoite-specific antigen SAG-2 but are different from the cysts formed by avirulentT. gondiistrains in that the internal parasite stages display ultrastructural features intermediate between tachyzoites and bradyzoites. Moreover, the zoites within the RH tissue cysts are clearly distinct from conventional bradyzoites in their sensitivity to pepsin-HCl digestion. In contrast to the observations made with partially resistant STAg/interleukin-12-vaccinated animals, no latent forms could be detected in brain tissue after RH challenge of mice immunized with a live attenuated tachyzoite vaccine which confers total protection against this parasite isolate. The above findings demonstrate the potential of a virulentT. gondiistrain to generate latent parasite stages, a process which may be promoted under conditions of incomplete vaccination.
Genomic Analysis of a Pathogenicity Island in Uropathogenic
Escherichia coli
CFT073: Distribution of Homologous Sequences among Isolates from Patients with Pyelonephritis, Cystitis, and CatheterAssociated Bacteriuria and from Fecal Samples
Authors: Guyer Debra M. , Kao Jyh-Shyang , Mobley Harry L. T.
Published: March 2020

Urinary tract infection is the most frequently diagnosed kidney and urologic disease and
Escherichia coli
is by far the most common etiologic agent. Uropathogenic strains have been shown to contain blocks of DNA termed pathogenicity islands (PAIs) which contribute to their virulence. We have defined one of these regions of DNA within the chromosome of a highly virulent
E. coli
strain, CFT073, isolated from the blood and urine of a woman with acute pyelonephritis. The 57,988-bp stretch of DNA has characteristics which define PAIs, including a size greater than 30 kb, the presence of insertion sequences, distinct segmentation of K-12 and J96 origin, GC content (42.9%) different from that of total genomic DNA (50.8%), and the presence of virulence genes (
hly
and
pap
). Within this region, we have identified 44 open reading frames; of these 44, 10 are homologous to entries in the complete K-12 genome sequence, 4 are nearly identical to the sequences of
E. coli
J96 encoding the HlyA hemolysin, 11 encode P fimbriae, and 19 show no homology to J96 or K-12 entries. To determine whether sequences found within the junctions of the PAI of CFT073 were common to other uropathogenic strains of
E. coli
, 11 probes were isolated along the length of the PAI and were hybridized to dot blots of genomic DNA isolated from clinical isolates (67 from patients with acute pyelonephritis, 38 from patients with cystitis, 49 from patients with catheter-associated bacteriuria, and 27 from fecal samples). These sequences were found significantly more often in strains associated with the clinical syndromes of acute pyelonephritis (79%) and cystitis (82%) than in those associated with catheter-associated bacteriuria (58%) and in fecal strains (22%) (
P
< 0.001). From these regions, we have identified a putative iron transport system and genes other than
hly
and
pap
that may contribute to the virulent phenotype of uropathogenic
E. coli
strains.
Early Events in Phagosome Establishment Are Required for Intracellular Survival ofLegionella pneumophila
Authors: Wiater Lawrence A. , Dunn Kenneth , Maxfield Frederick R. , Shuman Howard A.
Published: March 2020

During infection, the Legionnaires’ disease bacterium,Legionella pneumophila, survives and multiplies within a specialized phagosome that is near neutral pH and does not fuse with host lysosomes. In order to understand the molecular basis of this organism’s ability to control its intracellular fate, we have isolated and characterized a group of transposon-generated mutants which were unable to kill macrophages and were subsequently found to be defective in intracellular multiplication. These mutations define a set of 20 genes (19icm[for intracellular multiplication] genes anddotA[for defect in organelle trafficking]). In this report, we describe a quantitative assay for phagosome-lysosome fusion (PLF) and its use to measure the levels of PLF in cells that have been infected with either wild-typeL. pneumophilaor one of several mutants defective in differenticmgenes ordotA. By using quantitative confocal fluorescence microscopy, PLF could be scored on a per-bacterium basis by determining the extent to which fluorescein-labeledL. pneumophilacolocalized with host lysosomes prelabeled with rhodamine-dextran. Remarkably, mutations in the six genes that were studied resulted in maximal levels of PLF as quickly as 30 min following infection. These results indicate that several, and possibly all, of theicmanddotAgene products act at an early step during phagosome establishment to determine whetherL. pneumophila-containing phagosomes will fuse with lysosomes. Although not ruled out, subsequent activity of these gene products may not be necessary for successful intracellular replication.
Alteration of HLA-B27 Peptide Presentation after Infection of Transfected Murine L Cells byShigella flexneri
Authors: Boisgérault Florence , Mounier Joëlle , Tieng Vannary , Stolzenberg Marie-Claude , Khalil-Daher Iman , Schmid Michel , Sansonetti Philippe , Charron Dominique , Toubert Antoine
Published: March 2020

Shigella flexneriis a triggering agent for reactive arthritis in HLA-B27-susceptible individuals. Considering the intracellular multiplication of bacteria, it seems likely that bacterial peptides may be presented by the major histocompatibility complex (MHC) class I pathway. To examine this hypothesis, we infected HLA-B*2705- and/or human β2-microglobulin-transfected murine L-cell lines with M90T, an invasive strain ofS. flexneri. Bacterial infection induced no detectable modifications in the biosynthesis and expression level of HLA-B27, as assessed by immunoprecipitation, Northern blot analysis, and flow cytometry. Using confocal microscopy, we observed that bacterial infection induced a clustering of HLA-B27 molecules during macropinocytosis and before bacterial dissemination from cell to cell. Peptides naturally bound to HLA-B27 molecules were acid eluted from infected cells and separated by high-performance liquid chromatography. Major differences were observed in high-performance liquid chromatography profiles and in the nature of peptides presented following bacterial infection. Although most of the antigens presented were not accessed by Edman degradation, we obtained two sequences partially homologous to bacterial proteins. These peptides lacked the major HLA-B27 peptide anchor (Arg) at position 2, and one had an unusual length of 14 amino acids. These data suggest that alterations in the peptide presentation by HLA-B27 occur during infection, which could be relevant to the pathogenesis of HLA-B27-related arthritis.
Beta Interferon Is Produced by
Chlamydia trachomatis
-Infected Fibroblast-Like Synoviocytes and Inhibits Gamma Interferon-Induced HLA-DR Expression
Authors: Rödel Jürgen , Groh Annemarie , Vogelsang Heinz , Lehmann Marc , Hartmann Matthias , Straube Eberhard
Published: March 2020

Infection of fibroblast-like synovial cells with
Chlamydia trachomatis
(serotype D strain IC Cal 8) in culture induced the secretion of beta interferon (IFN-β). Chlamydial infection inhibited IFN-γ-induced expression of HLA-DR antigen in the cells. Addition of IFN-β antibody directly to infected cultures mitigated HLA-DR inhibition, suggesting involvement of produced IFN-β.
Synthesis and Characterization of a Protective Peptide-Based Vaccine againstSchistosoma mansoni
Authors: Tarrab-Hazdai Rebeca , Schechtman Deborah , Arnon Ruth
Published: March 2020

Two synthetic peptides, corresponding to the N-terminal sequence of the 45-kDa subunit of the protective 9B antigen ofSchistosoma mansoniand differing in only one amino acid residue, were synthesized. These peptides were recognized by the protective monoclonal antibody 152-66-9B, as well as by sera of mice and humans infected with schistosomiasis. The peptides were coupled to a protein carrier and used for immunization. One of the peptides, 9B-peptide1, induced in mice significant protection against challenge infection, manifested in a 40 to 50% reduction in worm burden.
The pH 6 Antigen of
Yersinia pestis
Binds to β1-Linked Galactosyl Residues in Glycosphingolipids
Authors: Payne Dean , Tatham David , Williamson E. Diane , Titball Richard W.
Published: March 2020

The
Yersinia pestis
pH 6 antigen was expressed by, and purified from,
Escherichia coli
containing cloned
psa
genes. By an enzyme-linked immunosorbence-based assay, purified pH 6 antigen bound to gangliotetraosylceramide (GM1A), gangliotriaosylceramide (GM2A), and lactosylceramide (LC) (designations follow the nomenclature of L. Svennerholm [J. Neurochem. 10:613–623, 1963]). Binding to GM1A, GM2A, and LC was saturable, with 50% maximal binding occurring at 498 ± 4, 390, and 196 ± 3 nM, respectively. Thin-layer chromatography (TLC) overlay binding confirmed that purified pH 6 antigen bound to GM1A, GM2A, and LC and also revealed binding to hydroxylated galactosylceramide. Intact
E. coli
cells which expressed the pH 6 antigen had a specificity similar to that of purified pH 6 in the TLC overlay assay except that nonhydroxylated galactosylceramide was also bound. The binding patterns observed indicate that the presence of β1-linked galactosyl residues in glycosphingolipids is the minimum determinant required for binding of the pH 6 antigen.
papG
Alleles among
Escherichia coli
Strains Causing Urosepsis: Associations with Other Bacterial Characteristics and Host Compromise
Authors: Johnson James R.
Published: March 2020

Alleles I, II, and III of the P adhesin gene
papG
were sought by PCR among 75
Escherichia coli
blood isolates from adults with urosepsis, and the
papG
genotype was compared with associated bacterial characteristics and host compromise status. Allele II predominated over allele III in the total population (71% of the strains versus 7% for allele III;
P
 < 0.01). Allele I was not encountered. In comparison with allele II, allele III was significantly associated with the presence of
hly
and the absence of
iuc
(which encode hemolysin and aerobactin biosynthesis), nonagglutination of digalactoside-coated beads, absence of aerobactin production, membership in serogroups O6 and O18, and host compromise, particularly cancer and upper urinary tract abnormalities.
The Alternative Sigma Factor, ς
E
, Is Critically Important for the Virulence of
Salmonella typhimurium
Authors: Humphreys Sue , Stevenson Andrew , Bacon Andrew , Weinhardt A. Barbara , Roberts Mark
Published: March 2020

In
Escherichia coli
, extracytoplasmic stress is partially controlled by the alternative sigma factor, RpoE (ς
E
). In response to environmental stress or alteration in the protein content of the cell envelope, ς
E
upregulates the expression of a number of genes, including
htrA
. It has been shown that
htrA
is required for intramacrophage survival and virulence in
Salmonella typhimurium
. To investigate whether ς
E
-regulated genes other than
htrA
are involved in salmonella virulence, we inactivated the
rpoE
gene of
S. typhimurium
SL1344 by allelic exchange and compared the phenotype of the mutant (GVB311) in vitro and in vivo with its parent and an isogenic
htrA
mutant (BRD915). Unlike
E. coli
, ς
E
is not required for the growth and survival of
S. typhimurium
at high temperatures. However, GVB311 did display a defect in its ability to utilize carbon sources other than glucose. GVB311 was more sensitive to hydrogen peroxide, superoxide, and antimicrobial peptides than SL1344 and BRD915. Although able to invade both macrophage and epithelial cell lines normally, the
rpoE
mutant was defective in its ability to survive and proliferate in both cell lines. The effect of the
rpoE
mutation on the intracellular behavior of
S. typhimurium
was greater than that of the
htrA
mutation. Both GVB311 and BRD915 were highly attenuated in mice. Neither strain was able to kill mice via the oral route, and the 50% lethal dose (LD
50
) for both strains via the intravenous (i.v.) route was very high. The i.v. LD
50
s for SL1344, BRD915, and GVB311 were <10, 5.5 × 10
5
, and 1.24 × 10
7
CFU, respectively. Growth in murine tissues after oral and i.v. inoculation was impaired for both the
htrA
and
rpoE
mutant, with the latter mutant being more severely affected. Neither mutant was able to translocate successfully from the Peyer’s patches to other organs after oral infection or to proliferate in the liver and spleen after i.v. inoculation. However, the
htrA
mutant efficiently colonized the livers and spleens of mice infected i.v., but the
rpoE
mutant did not. Previous studies have shown that salmonella
htrA
mutants are excellent live vaccines. In contrast, oral immunization of mice with GVB311 was unable to protect any of the mice from oral challenge with SL1344. Furthermore, i.v. immunization with a large dose (∼10
6
CFU) of GVB311 protected less than half of the orally challenged mice. Thus, our results indicate that genes in the ς
E
regulon other than
htrA
play a critical role in the virulence and immunogenicity of
S. typhimurium
.
Immunological Characterization of a Protective Antigen of
Erysipelothrix rhusiopathiae
: Identification of the Region Responsible for Protective Immunity
Authors: Shimoji Yoshihiro , Mori Yasuyuki , Fischetti Vincent A.
Published: March 2020

The gene encoding a protective protein antigen of the gram-positive bacterium
Erysipelothrix rhusiopathiae
, an important veterinary pathogen responsible for erysipelas in swine and a variety of diseases in animals, was cloned and sequenced. The gene encodes a polypeptide of 597 amino acids plus a putative signal sequence of 29 amino acids, resulting in a mature protein with a molecular mass of 69,017 Da. Sequence analysis of the gene product revealed a C-terminal region composed of nine tandem repeats of 20 amino acids and a total sequence that is nearly identical to that of the 64-kDa cell surface protein (SpaA) of the bacterium. Because of this similarity, the protein was designated SpaA.1. In this study, we examined whether the SpaA.1 protein could induce protective antibodies and whether we could identify the region involved in protective immunity. Both the mature SpaA.1 protein and its C-terminal repeat region, but not the N-terminal segment, were expressed in
Escherichia coli
and purified as a histidine-tagged fusion recombinant protein. Rabbit antiserum raised against the mature SpaA.1 protein passively protected mice from lethal challenge with a virulent homologous strain, Fujisawa-SmR, suggesting that protection is mediated by humoral antibodies. To determine which domain of the SpaA.1 protein is responsible for the observed protection, mice were actively immunized with either the mature SpaA.1 protein or the C-terminal repeat region and then challenged with Fujisawa-SmR. The result showed that mice immunized with the mature SpaA.1 protein, but not the C-terminal repeat region, were protected, suggesting that the protection-eliciting epitope(s) is located within the N-terminal two-thirds of the SpaA.1 molecule. This was confirmed by passive immunization experiments in which the protective activity of rabbit antiserum, raised against mature SpaA.1 protein, was not abolished by absorption with the purified recombinant C-terminal repeat region. In addition, antibodies specific for the C-terminal repeat region were unable to protect mice from lethal challenge. These results show that the N-terminal two-thirds of the SpaA.1 molecule may constitute a good vaccine candidate against erysipelas.
Construction and Characterization of Mutations within the
Klebsiella mrkD
1P
Gene That Affect Binding to Collagen Type V
Authors: Sebghati Tricia A. , Clegg Steven
Published: March 2020

The fimbria-associated MrkD
1P
protein mediates adherence of type 3 fimbriate strains of
Klebsiella pneumoniae
to collagen type V. Currently, three different MrkD adhesins have been described in
Klebsiella
species, and each possesses a distinctive binding pattern. Therefore, the binding abilities of mutants possessing defined mutations within the
mrkD
1P
gene were examined in order to determine whether specific regions of the adhesin molecule were responsible for collagen binding. Both site-directed and chemically induced mutations were constructed within
mrkD
1P
, and the ability of the gene products to be incorporated into fimbrial appendages or bind to collagen was determined. Binding to type V collagen was not associated solely with one particular region of the MrkD
1P
protein, and two classes of nonadhesive mutants were isolated. In one class of mutants, the MrkD adhesin was not assembled into the fimbrial shaft, whereas in the second class of mutants, the adhesin was associated with fimbriae but did not bind to collagen. Both hemagglutinating and collagen-binding activities were associated with the MrkD
1P
molecule, since P pili and type 3 fimbriae carrying adhesive MrkD proteins exhibited identical binding properties.