bluuebunny/crossref_metadata_2025_split huggingface dataset:
Comparison of Surface Proteins ofAnaplasma marginaleGrown in Tick Cell Culture, Tick Salivary Glands, and Cattle
Authors: Barbet A. F. , Blentlinger R. , Yi Jooyoung , Lundgren A. M. , Blouin E. F. , Kocan K. M. 
Published: December 2019

ABSTRACTAnaplasma marginale, a tick-borne rickettsial pathogen of cattle, infects bovine erythrocytes, resulting in mild to severe hemolytic disease that causes economic losses in domestic livestock worldwide. Recently, the Virginia isolate ofA. marginalewas propagated in a continuous tick cell line, IDE8, derived from embryonicIxodes scapularis. Development ofA. marginalein cell culture was morphologically similar to that described previously in ticks. In order to evaluate the potential of the cell culture-derived organisms for use in future research or as an antigen for serologic tests and vaccines, the extent of structural conservation of the major surface proteins (MSPs) between the cell culture-derivedA. marginaleand the bovine erythrocytic stage, currently the source ofA. marginaleantigen, was determined. Structural conservation on the tick salivary-gland stage was also examined. Monoclonal and monospecific antisera against MSPs 1 through 5, initially characterized against erythrocyte stages, also reacted withA. marginalefrom cell culture and tick salivary glands. MSP1a among geographicA. marginaleisolates is variable in size because of different numbers of a tandemly repeated 28- or 29-amino-acid peptide. The cell culture-derivedA. marginalemaintained the same-size MSP1a as that found on the Virginia isolate ofA. marginalein bovine erythrocytes and tick salivary glands. Although differences were observed in the polymorphic MSP2 antigen between culture and salivary-gland stages, MSP2 did not appear to vary, by two-dimensional gel electrophoresis, during continuous passage in culture. These data show that MSPs of erythrocyte-stageA. marginaleare present on culture stages and may be structurally conserved during continuous culture. The presence of all current candidate diagnostic and vaccine antigens suggests that in vitro cultures are a valuable source of rickettsiae for basic research and for the development of improved diagnostic reagents and vaccines against anaplasmosis.
Expression of the EspB Protein of Enteropathogenic
Escherichia coli
within HeLa Cells Affects Stress Fibers and Cellular Morphology
Authors: Taylor Kathleen A. , Luther Paul W. , Donnenberg Michael S. 
Published: December 2019

ABSTRACT

The EspB protein of enteropathogenic
Escherichia coli
(EPEC) is essential for the signaling events that lead to the accumulation of actin beneath intimately attached bacteria, a process that is known as the attaching and effacing effect. EspB is targeted to the host cell cytoplasm by a type III secretion apparatus. To determine the effect of intracellular EspB on the host cell cytoskeleton, we transfected HeLa cells with a plasmid containing the
espB
gene under the control of an inducible eukaryotic promoter. A HeLa cell clone that expressed
espB
mRNA and EspB protein after induction was selected for further study. The expression of EspB in these cells caused a dramatic change in cell morphology and a marked reduction in actin stress fibers. Cells expressing EspB were significantly impaired in their ability to support invasion by EPEC and
Salmonella typhimurium
. However, the expression of EspB within host cells could not compensate for the lack of EspB expression by an
espB
mutant strain of EPEC to restore attaching and effacing activity. These studies suggest that EspB is a cytoskeletal toxin that is translocated to the host cell cytoplasm, where it causes a redistribution of actin.
The Immunoglobulin (IgG) Antibody Response to OspA and OspB Correlates with Severe and Prolonged Lyme Arthritis and the IgG Response to P35 Correlates with Mild and Brief Arthritis
Authors: Akin Evren , McHugh Gail L. , Flavell Richard A. , Fikrig Erol , Steere Allen C. 
Published: December 2019

ABSTRACTIn an effort to implicate immune responses to specificBorrelia burgdorferiproteins that may have a role in chronic Lyme arthritis, we studied the natural history of the antibody response toB. burgdorferiin serial serum samples from 25 patients monitored throughout the course of Lyme disease. In these patients, the immunoglobulin G (IgM) and IgG antibody responses to 10 recombinantB. burgdorferiproteins, determined during early infection, early arthritis, and maximal arthritis, were correlated with the severity and duration of maximal arthritis. The earliest responses were usually to outer surface protein C (OspC), P35, P37, and P41; reactivity with OspE, OspF, P39, and P93 often developed weeks later; and months to years later, 64% of patients had responses to OspA and OspB. During early infection and early arthritis, the levels of IgG antibody to P35 correlated inversely with the subsequent severity or duration of maximal arthritis. In contrast, during periods of maximal arthritis, the levels of IgG antibody to OspA and OspB, especially to a C-terminal epitope of OspA, correlated directly with the severity and duration of arthritis. Thus, the higher the IgG antibody response to P35 earlier in the infection, the milder and briefer the subsequent arthritis, whereas during maximal arthritis, the higher the IgG response to OspA and OspB, the more severe and prolonged the arthritis.
Mechanisms for Induction of L-Selectin Loss from T Lymphocytes by a Cryptococcal Polysaccharide, Glucuronoxylomannan
Authors: Dong Zhao Ming , Jackson Lydgia , Murphy Juneann W. 
Published: December 2019

ABSTRACTDisseminated cryptococcosis is accompanied by cryptococcal polysaccharides in the serum and the lack of cellular infiltrates in infected tissues. Cryptococcal polysaccharides given intravenously to mice inhibit the influx of T lymphocytes into the sites of cell-mediated immune response. The focus here was to determine whether cryptococcal polysaccharides modulate the expression of molecules, such as L-selectin, that are important in extravasation of T cells. Cryptococcal glucuronoxylomannan (GXM), but not galactoxylomannan or mannoprotein, was found to cause loss of L-selectin from freshly isolated human T cells of both CD4 and CD8 subsets and from Jurkat cells. With the signaling-pathway inhibitors staurosporine (which inhibits protein kinase C) and herbimycin A (which inhibits protein tyrosine kinases), we showed that GXM or the cryptococcal culture filtrate antigen CneF directly induces L-selectin loss from CD4+and CD8+T cells via a herbimycin A-sensitive pathway(s) presumably involving one or more protein tyrosine kinases but not via a pathway involving protein kinase C. Loss of L-selectin from the T cells before the T cells have a chance to bind to L-selectin ligands on endothelial cells would be expected to prevent T-cell migration into inflamed tissues and/or lymph organs.
Existing Antilisterial Immunity Does Not Inhibit the Development of aListeria monocytogenes-Specific Primary Cytotoxic T-Lymphocyte Response
Authors: Bouwer H. G. Archie , Shen Hao , Fan Xin , Miller Jeff F. , Barry Ronald A. , Hinrichs David J. 
Published: December 2019

ABSTRACTInfection of BALB/c mice withListeria monocytogenesstimulates an antilisterial immune response evident by the appearance of H2-Kd-restricted CD8+cytotoxic T lymphocytes (CTLs) specific for the nanomer peptides amino acids (aa) 91 to 99 of listeriolysin O (LLO 91–99) and aa 217 to 225 of the p60 molecule (p60 217–225). We have introduced point mutations at anchor residues within LLO 91–99 (92F) or p60 217–225 (218F), and BALB/c mice infected withL. monocytogenesstrains containing these point mutations do not develop CTLs specific for LLO 91–99 or p60 217–225, respectively. We have used these strains to test whether primary CTL responses againstL. monocytogenes-derived determinants can be stimulated within an environment of existing antilisterial immunity. We found that the development of a primaryL. monocytogenes-specific CTL response is not altered by existing immunity toL. monocytogenes. For example, primary immunization with the p60 218F strain ofL. monocytogenesfollowed by a secondary immunization with wild-typeL. monocytogenesresults in stimulation of p60 217–225-specific CTLs at primary response levels and LLO 91–99-specific effectors at levels consistent with a memory CTL response. Similarly, primary immunization with the 92F strain ofL. monocytogenesfollowed by a secondary immunization with wild-typeL. monocytogenesresults in stimulation of LLO 91–99-specific CTLs at primary response levels and p60 217–225-specific effectors at levels consistent with a memory CTL response. These results provide additional support for the use ofL. monocytogenesas a recombinant vaccine vector and show that antivector immunity does not inhibit the development of a primary CTL response when the epitope is delivered byL. monocytogenesas the vaccine strain.
Establishment of Unipolar Localization of IcsA in
Shigella flexneri
2a Is Not Dependent on Virulence Plasmid Determinants
Authors: Sandlin Robin C. , Maurelli Anthony T. 
Published: December 2019

ABSTRACT

Unipolar localization of IcsA on the surface of
Shigella flexneri
is required for efficient formation of actin tails and protrusions in infected eucaryotic cells. Lipopolysaccharide (LPS) mutations have been demonstrated to affect either the establishment or the maintenance of IcsA in a unipolar location, although the mechanism is unknown. In order to analyze the contribution of virulence plasmid determinants on the unipolar localization of IcsA, we examined the localization of IcsA expressed from a cloned plasmid copy in two different genetic backgrounds. The localization of IcsA was first examined in a virulence plasmid-cured derivative of the wild-type
S. flexneri
2a isolate 2457T. This approach examined the contribution of virulence plasmid-borne factors, including the previously identified virulence plasmid-borne protease that is responsible for cleaving IcsA in the outer membrane and releasing the 95-kDa secreted form from the cell surface. IcsA localization in a related but nonpathogenic
Escherichia coli
strain expressing LPS of the O8 serotype was also examined. IcsA surface presentation in both of these genetic backgrounds continued to be unipolar, demonstrating that virulence plasmid-borne determinants are not responsible for unipolar localization of IcsA. The unipolar localization of IcsA in the
E. coli
background suggests that a common pathway that allows IcsA to be spatially restricted to one pole on the bacterial cell surface exists in
Shigella
and
E. coli
.
Recombinant Expression and Localization of
Schistosoma mansoni
Cathepsin L1 Support Its Role in the Degradation of Host Hemoglobin
Authors: Brady Ciaran P. , Dowd Andrew J. , Brindley Paul J. , Ryan Thecla , Day Sharon R. , Dalton John P. 
Published: December 2019

ABSTRACT

Cysteine proteinases expressed by schistosomes appear to play key roles in the digestion of host hemoglobin, the principal source of amino acid nutrients utilized by these parasites. We have shown previously that the predominant cysteine proteinase activity in soluble extracts and excretory/secretory (ES) products of adults of
Schistosoma mansoni
and
S. japonicum
is cathepsin L-like in its substrate specificity. However, biochemical analysis of the cathepsin L activity in extracts and ES products of schistosomes has been complicated by the presence of at least two distinct forms of schistosome cathepsin L, termed SmCL1 and SmCL2. We now report the purification and enzyme characteristics of active, recombinant SmCL1 which was obtained by transforming
Saccharomyces cerevisiae
with an expression plasmid encoding the preproenzyme of SmCL1. Recombinant SmCL1 was secreted by the transformed yeast into the culture media from which it was purified by gel filtration and ion-exchange chromatography. The purified enzyme exhibited substrate specificity against synthetic peptidyl substrates (e.g., Boc-Val-Leu-Lys-NHMec and Z-Phe-Arg-NHMec;
k
cat
/

K
m

= 17.25 and 6.24 mM
−1
s
−1
, respectively) and against gelatin and hemoglobin, characteristic of cathepsin L. Immunoblot analysis using antiserum raised against recombinant SmCL1 demonstrated that native SmCL1 of 33 kDa was present in ES products and soluble extracts of
S. mansoni
. Using this antiserum and thin tissue sections, we localized the native SmCL1 to the gastrodermis and to the tegument of adult schistosomes. Recombinant SmCL1 was capable of degrading human hemoglobin at pH 4.0 to 4.5 but not higher, suggesting that denaturation of hemoglobin by low pH, as found in the cecum of the adult schistosome, may be necessary for its catalysis by cathepsin L and other gut-associated proteinases. Together, these results support a role for SmCL1 in the degradation of host hemoglobin within the gut of the schistosome.
Tonsillar Application of Formalin-Killed Cells ofStreptococcus sobrinusReduces Experimental Dental Caries in Rabbits
Authors: Fukuizumi Takaki , Inoue Hiromasa , Tsujisawa Toshiyuki , Uchiyama Choji 
Published: December 2019

ABSTRACTLivingStreptococcus sobrinuscells were orally inoculated into nonimmune rabbits and rabbits immunized with formalin-killed cells ofS. sobrinusthrough tonsillar application to examine the anticaries potential of this method of immunization. The livingS. sobrinuscell numbers and the caries areas in the rabbits immunized by tonsillar application decreased to a level one-fifth of that in nonimmune rabbits.
High-Level Expression of
Plasmodium vivax
Apical Membrane Antigen 1 (AMA-1) in
Pichia pastoris
: Strong Immunogenicity in
Macaca mulatta
Immunized with
P. vivax
AMA-1 and Adjuvant SBAS2
Authors: Kocken Clemens H. M. , Dubbeld Martin A. , Van Der Wel Annemarie , Pronk Jack T. , Waters Andrew P. , Langermans Jan A. M. , Thomas Alan W. 
Published: December 2019

ABSTRACT

The apical membrane antigen 1 (AMA-1) family is a promising family of malaria blood-stage vaccine candidates that have induced protection in rodent and nonhuman primate models of malaria. Correct conformation of the protein appears to be essential for the induction of parasite-inhibitory responses, and these responses appear to be primarily antibody mediated. Here we describe for the first time high-level secreted expression (over 50 mg/liter) of the
Plasmodium vivax
AMA-1 (PV66/AMA-1) ectodomain by using the methylotrophic yeast
Pichia pastoris
. To prevent nonnative glycosylation, a conservatively mutagenized PV66/AMA-1 gene (PV66Δglyc) lacking N-glycosylation sites was also developed. Expression of the PV66Δglyc ectodomain yielded similar levels of a homogeneous product that was nonglycosylated and was readily purified by ion-exchange and gel filtration chromatographies. Recombinant PV66Δglyc
43–487
was reactive with conformation-dependent monoclonal antibodies. With the SBAS2 adjuvant,
Pichia
-expressed PV66Δglyc
43–487
was highly immunogenic in five rhesus monkeys, inducing immunoglobulin G enzyme-linked immunosorbent assay titers in excess of 1:200,000. This group of monkeys had a weak trend showing lower cumulative parasite loads following a
Plasmodium cynomolgi
infection than in the control group.
Killing of
Borrelia burgdorferi
by Antibody Elicited by OspA Vaccine Is Inefficient in the Absence of Complement
Authors: Nowling Jena M. , Philipp Mario T. 
Published: December 2019

ABSTRACT

A Lyme disease vaccine, based on the
Borrelia burgdorferi
lipoprotein OspA, has recently undergone phase III trials in humans. The results of one of these trials indicate that vaccine efficacy positively correlates with anti-OspA antibody titer. Spirochete killing within the tick vector midgut, upon which vaccine efficacy appears to depend, may occur chiefly via a mechanism that involves antibody alone, as it has been reported that complement is degraded by tick saliva decomplementing factors. We compared the in vitro killing efficiencies of anti-OspA antibody elicited in rhesus monkeys by the OspA vaccine, in the presence and in the absence of monkey complement. Killing in the absence of complement was between 14 and 3,800 times less efficient than with complement present, depending on the spirochete strain. The relative inefficiency of the complement-independent killing mechanism by anti-OspA antibody may explain why OspA vaccine efficacy is critically dependent on antibody titer.
Murine γδ T Lymphocytes Elicited duringPlasmodium yoeliiInfection Respond toPlasmodiumHeat Shock Proteins
Authors: Kopacz Jeffrey , Kumar Nirbhay 
Published: December 2019

ABSTRACTγδ T cells accumulate duringPlasmodiuminfections in both murine and human malarias. The biological role of these cells and the antigens that they recognize are not clearly understood, although recent findings indicate that γδ T cells in general influence both innate and antigen-specific adaptive host responses. We examined the accumulation of γδ T cells elicited during infection with virulent and avirulentPlasmodium yoeliiparasites in relatively susceptible and resistant strains of mice. Our results indicated that in nonlethal malaria infections, γδ T cells comprise a larger proportion of splenic T cells than in lethal infections and that only a live infection is capable of inducing an increase in the percentage of γδ T cells in vivo. Furthermore, we demonstrate that γδ T cells elicited during aP. yoeliiinfection respond by proliferation in vitro toP. falciparumheat shock proteins (HSPs) of 60 and 70 kDa, suggesting a possible immunological involvement of parasite HSPs in this arm of the cellular immune response during malarial infection in mice.
Campylobacter jejuni
-Stimulated Secretion of Interleukin-8 by INT407 Cells
Authors: Hickey Thomas E. , Baqar Shihida , Bourgeois A. Louis , Ewing Cheryl P. , Guerry Patricia 
Published: December 2019

ABSTRACT

Incubation of INT407 cells with various clinical isolates of
Campylobacter jejuni
resulted in secretion of interleukin-8 (IL-8) at levels ranging from 96 to 554 pg/ml at 24 h. The strains which produced the highest levels of IL-8 secretion were 81-176 and BT44. Induction of IL-8 secretion required live cells of 81-176 and was dependent on de novo protein synthesis. Site-specific mutants of 81-176, which were previously shown to be defective in adherence and invasion, resulted in reduced levels of secretion of IL-8, and
cheY
mutants of strains 81-176 and 749, which are hyperadherent and hyperinvasive, resulted in higher levels of IL-8 secretion. Another mutant of 81-176, which adheres at about 43% of the wild-type levels but is noninvasive, also showed marked reduction in IL-8 levels, suggesting that invasion is necessary for high levels of IL-8 secretion. When gentamicin was added to INT407 cells at 2 h after infection with 81-176, IL-8 secretion 22 h later was equivalent to that of controls without gentamicin, suggesting that the events which trigger induction and release of IL-8 occur early in the interactions of bacteria and eukaryotic cells.
Roles of Glycoproteins and Oligosaccharides Found in Human Vaginal Fluid in Bacterial Adherence
Authors: Rajan Nithya , Cao Qing , Anderson Byron E. , Pruden Denise L. , Sensibar Julia , Duncan James L. , Schaeffer Anthony J. 
Published: December 2019

ABSTRACTAdherence of type 1-piliatedEscherichia colito carbohydrate structures of vaginal mucosa plays a major role in the pathogenesis of ascending urinary tract infections in women. Colonization of the vaginal introitus is influenced by interactions between pathogens, vaginal fluid, and vaginal epithelium. In this study, the type and amount of carbohydrates and glycoproteins present in vaginal fluid were determined. Free and protein-bound oligosaccharides in vaginal fluid specimens were analyzed by fluorophore-assisted carbohydrate electrophoresis (FACE) and high-pressure liquid chromatography (HPLC). Two-dimensional electrophoretic separations of vaginal fluid glycoproteins were performed together with bacterial overlay assays. The results of FACE showed that the majority of the oligosaccharides are in the free state and the bound oligosaccharides are undetectable. HPLC analysis of free sugars revealed glucose as the major sugar (3.3 ± 0.3 mM), and the concentrations of mannose and glucosamine were 0.065 ± 0.04 and 0.02 ± 0.001 mM, respectively. RadiolabeledE. colibound three vaginal fluid glycoproteins with the following molecular masses and pIs: 82 kDa and pI 5.5, 55 kDa and pI 4.5, and 55 kDa and pI 6.5. The binding was inhibited by mannose and by deglycosylation of the proteins prior to the overlay assay. One of these putative receptors was identified to be the heavy chain of secretory IgA (S-IgA). These data suggest that the free mannose in the fluid is less than that required to affectE. coli-epithelial cell binding interactions and that S-IgA may bindE. coliin the vaginal introitus.
Humoral and Cellular Immune Responses in Mice Immunized with RecombinantMycobacterium bovisBacillus Calmette-Guérin Producing a Pertussis Toxin-Tetanus Toxin Hybrid Protein
Authors: Abomoelak B. , Huygen K. , Kremer L. , Turneer M. , Locht C. 
Published: December 2019

ABSTRACTThe development of combined vaccines constitutes one of the priorities in modern vaccine research. One of the most successful combined vaccines in use is the diphtheria-pertussis-tetanus vaccine. However, concerns about the safety of the pertussis arm have led to decreased acceptance of the vaccine but also to the development of new, safer, and effective acellular vaccines against pertussis. Unfortunately, the production cost of these new vaccines is significantly higher than that of previous vaccines. Here, we explore the potential of live recombinantMycobacterium bovisBCG producing the hybrid protein S1-TTC, which contains the S1 subunit of pertussis toxin fused to fragment C of tetanus toxin, as an alternative to the acellular vaccines. S1-TTC was produced in two different expression systems. In the first system its production was under the control of the 85A antigen promoter and signal peptide, and in the second system it was under the control of thehsp60promoter. Although expression of the hybrid antigen was obtained in both cases, only the second expression system yielded a recombinant BCG strain able to induce both a specific humoral immune response and a specific cellular immune response. The antibodies generated were directed against the TTC part and neutralized toxin activity in an in vivo challenge model, whereas interleukin-2 production was specific for both parts of the molecule. Since protection against tetanus is antibody mediated and protection against pertussis may be cell mediated, this constitutes a first promising step towards the development of a cost-effective, protective, and safe combined vaccine against pertussis, tetanus, and tuberculosis.