bluuebunny/crossref_metadata_2025 huggingface dataset:
Prior Immunity to Homologous and Heterologous
Salmonella
Serotypes Suppresses Local and Systemic Anti-Fragment C Antibody Responses and Protection from Tetanus Toxin in Mice Immunized with
Salmonella
Strains Expressing Fragment C
Authors: Roberts Mark , Bacon Andrew , Li Jingli , Chatfield Steven
Published: December 2019

We have investigated the effect of preexisting immunity to homologous (
Salmonella typhimurium
) or heterologous (
S. dublin
) serotypes of
Salmonella
on the ability of an attenuated
S. typhimurium aroA aroD
vector (BRD509) to immunize mice against the heterologous antigen fragment C (FrgC). We studied two strains, BRD847 and BRD937, expressing FrgC carried on plasmids that differ only with respect to the promoter controlling FrgC expression, the
nirB
promoter in the case of BRD847 and the
htrA
promoter in the case of BRD937. Mice were preimmunized orally with
S. typhimurium
BRD509,
S. dublin aroA aroD
(BRD620), or saline. Forty-four days later, they were immunized orally with BRD847 or BRD937. Prior immunity to
S. typhimurium
severely depressed the serum immunoglobulin G (IgG) and IgA anti-FrgC response in both BRD847- and BRD937-immunized mice. Mice with existing immunity to
S. dublin
also had lower IgG anti-FrgC geometric mean titers (GMTs) than did mice preimmunized with saline, but this difference was significant only in the case of mice immunized with BRD937. However, in nonimmune mice or in mice preimmunized with
S. typhimurium
or
S. dublin
, the anti-FrgC IgG GMTs were always higher in mice in the BRD937 groups than in the equivalent BRD847 groups. This is reflected in the effect of prior immunity on the ability of oral immunization with BRD847 or BRD937 to protect mice from challenge with a lethal dose of tetanus toxin. All of the mice preimmunized with saline and then immunized with BRD847 or BRD937 survived challenge. Only 20% of the animals immunized with BRD847 and 60% of the mice in the BRD937 group survived tetanus toxin challenge if they were preimmunized with BRD509. Preexisting immunity to
S. dublin
did not affect the ability of BRD937 to immunize mice against tetanus, but it did reduce the efficiency of BRD847: only 60% percent of the mice survived challenge. The intestinal secretory IgA responses to FrgC were very similar in the BRD847 and BRD937 groups. Prior immunity did depress the IgA anti-FrgC titers but only significantly so in the mice preimmunized with BRD509. These results show that preexisting
Salmonella
immunity, particularly to homologous serotypes, can severely compromise the ability of live
Salmonella
vectors to deliver heterologous antigens to the mammalian immune system. However, the results also indicate that this may be overcome by the design of more powerful in vivo expression systems.
Selective Distribution of a High-Affinity Plasminogen-Binding Site among Group A Streptococci Associated with Impetigo
Authors: Svensson Mikael D. , Sjöbring Ulf , Bessen Debra E.
Published: December 2019

Group A streptococci can be classified according to their tendency to cause either impetigo, pharyngitis, or both types of infection. Genotypic markers for tissue site preference lie withinemmgenes, which encode fibrillar surface proteins that play a key role in virulence.emmgene products (M and M-like proteins) display an extensive array of binding activities for tissue and plasma proteins of the human host. In a previous study, a high-affinity binding site for human plasmin(ogen) was mapped to theemm53gene product. In this report, a structurally similar plasminogen-binding domain is found to be widely and selectively distributed among group A streptococci harboring theemmgene marker for the skin as the preferred tissue site for infection. The findings are highly suggestive of a central role for bacterial modulation of host plasmin(ogen) during localized infection at the epidermis.
Cytoadherence of
Babesia bovis
-Infected Erythrocytes to Bovine Brain Capillary Endothelial Cells Provides an In Vitro Model for Sequestration
Authors: O’Connor Roberta M. , Long Jennifer A. , Allred David R.
Published: December 2019

Babesia bovis
, an intraerythrocytic parasite of cattle, is sequestered in the host microvasculature, a behavior associated with cerebral and vascular complications of this disease. Despite the importance of this behavior to disease etiology, the underlying mechanisms have not yet been investigated. To study the components involved in sequestration,
B. bovis
parasites that induce adhesion of the infected erythrocytes (IRBCs) to bovine brain capillary endothelial cells (BBEC) in vitro were isolated. Two clonal lines, CD7
A+I+
and CE11
A+I−
, were derived from a cytoadherent, monoclonal antibody 4D9.1G1-reactive parasite population. This antibody recognizes a variant, surface-exposed epitope of the variant erythrocyte surface antigen 1 (VESA1) of
B. bovis
IRBCs. Both clonal lines were cytoadhesive to BBEC and two other bovine endothelial cell lines but not to COS7 cells, FBK-4 cells, C32 melanoma cells, or bovine brain pericytes. By transmission electron microscopy, IRBCs were observed to bind to BBEC via the knobby protrusions on the IRBC surface, indicating involvement of components associated with these structures. Inhibition of protein export in intact, trypsinized IRBCs ablated both erythrocyte surface reexpression of parasite protein and cytoadhesion. IRBCs allowed to recover surface antigen expression regained the ability to bind endothelial cells, demonstrating that parasite protein export is required for cytoadhesion. We propose the use of this assay as an in vitro model to study the components involved in
B. bovis
cytoadherence and sequestration.
A Random Survey of the
Cryptosporidium parvum
Genome
Authors: Liu Chang , Vigdorovich Vladimir , Kapur Vivek , Abrahamsen Mitchell S.
Published: December 2019

Cryptosporidium parvum
is an obligate intracellular pathogen responsible for widespread infections in humans and animals. The inability to obtain purified samples of this organism’s various developmental stages has limited the understanding of the biochemical mechanisms important for
C. parvum
development or host-parasite interaction. To identify
C. parvum
genes independent of their developmental expression, a random sequence analysis of the 10.4-megabase genome of
C. parvum
was undertaken. Total genomic DNA was sheared by nebulization, and fragments between 800 and 1,500 bp were gel purified and cloned into a plasmid vector. A total of 442 clones were randomly selected and subjected to automated sequencing by using one or two primers flanking the cloning site. In this way, 654 genomic survey sequences (GSSs) were generated, corresponding to >320 kb of genomic sequence. These sequences were assembled into 408 contigs containing >250 kb of unique sequence, representing ∼2.5% of the
C. parvum
genome. Comparison of the GSSs with sequences in the public DNA and protein databases revealed that 107 contigs (26%) displayed similarity to previously identified proteins and rRNA and tRNA genes. These included putative genes involved in the glycolytic pathway, DNA, RNA, and protein metabolism, and signal transduction pathways. The repetitive sequence elements identified included a telomere-like sequence containing hexamer repeats, 57 microsatellite-like elements composed of dinucleotide or trinucleotide repeats, and a direct repeat sequence. This study demonstrates that large-scale genomic sequencing is an efficient approach to analyze the organizational characteristics and information content of the
C. parvum
genome.
Thymic Independence of Adaptive Immunity to the Intracellular PathogenShigella flexneriSerotype 2a
Authors: Way Sing Sing , Borczuk Alain C. , Goldberg Marcia B.
Published: December 2019

Shigella flexneriis a facultative intracellular pathogen. While immunity to several intracellular pathogens is mediated by T lymphocytes, it is unknown whether cellular immune responses are important to adaptive immunity toS. flexneri. We show that vaccination withS. flexneriserotype 2a confers protection to mice that lack T lymphocytes or gamma interferon (IFN-γ), specific depletion of T lymphocytes does not alter the protection, and adoptive transfer of splenocytes from vaccinated mice does not confer protection to naive mice. In contrast, vaccination conferred no protection to mice that lack B lymphocytes and adoptive transfer of immune sera conferred partial protection to naive mice. These data demonstrate that in the mouse bronchopulmonary model, adaptive immunity toS. flexneri2a is an antibody-mediated, B-lymphocyte-dependent process and can be generated in the absence of T lymphocytes or IFN-γ.
New Exfoliative Toxin Produced by a Plasmid-Carrying Strain of
Staphylococcus hyicus
Authors: Sato Hisaaki , Watanabe Takao , Murata Yasuko , Ohtake Ayumi , Nakamura Mayumi , Aizawa Chikara , Saito Hiroshi , Maehara Nobutoshi
Published: December 2019

A new serotype of
Staphylococcus hyicus
exfoliative toxin (SHET), serotype B, was isolated from the culture filtrate of a plasmid-carrying strain of
S. hyicus
. The new SHET was purified by precipitation with 70% saturated ammonium sulfate, gel filtration on a Sephadex G-75 column, column chromatography on DEAE–Cellulofine A-500, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The new SHET caused exfoliation of the epidermis as determined by the so-called Nikolsky sign when inoculated into 1-day-old chickens. The new SHET was serologically different from
Staphylococcus aureus
exfoliative toxins (ETs) (ETA, ETB, and ETC) and from the SHET from the plasmidless strain but showed the same molecular weight as the other serotypes of toxins on SDS-PAGE. It was thermolabile and lost its toxicity after being heated at 60°C for 30 min. We propose that the new SHET be designated SHETB and that the SHET produced by the plasmidless strain be designated SHETA.
Characterization of an Enterotoxigenic
Escherichia coli
Strain from Africa Expressing a Putative Colonization Factor
Authors: Khalil Sami B. , Cassels Frederick J. , Shaheen Hind I. , Pannell Lewis K. , El-Ghorab Nemat , Kamal Karim , Mansour Moustafa , Savarino Stephen J. , Peruski Leonard F.
Published: December 2019

An enterotoxigenic
Escherichia coli
(ETEC) strain of serotype O114:H− that expressed both heat-labile and heat-stable enterotoxins and tested negative for colonization factors (CF) was isolated from a child with diarrhea in Egypt. This strain, WS0115A, induced hemagglutination of bovine erythrocytes and adhered to the enterocyte-like cell line Caco-2, suggesting that it may elaborate novel fimbriae. Surface-expressed antigen purified by differential ammonium sulfate precipitation and column chromatography yielded a single protein band with
M
r
14,800 when resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (16% polyacrylamide). A monoclonal antibody against this putative fimbrial antigen was generated and reacted with strain WS0115A and also with CS1-, CS17-, and CS19-positive strains in a dot blot assay. Reactivity was temperature dependent, with cells displaying reactivity when grown at 37°C but not when grown at 22°C. Immunoblot analysis of a fimbrial preparation from strain WS0115A showed that the monoclonal antibody reacted with a single protein band. Electron microscopy and immunoelectron microscopy revealed fimbria-like structures on the surface of strain WS0115A. These structures were rigid and measured 6.8 to 7.4 nm in diameter. Electrospray mass-spectrometric analysis showed that the mass of the purified fimbria was 14,965 Da. The N-terminal sequence of the fimbria established that it was a member of the CFA/I family, with sequence identity to the amino terminus of CS19, a new CF recently identified in India. Cumulatively, our results suggest that this fimbria is CS19. Screening of a collection of ETEC strains isolated from children with diarrhea in Egypt found that 4.2% of strains originally reported as CF negative were positive for this CF, suggesting that it is biologically relevant in the pathogenesis of ETEC.
A Murine Model for Infection withBurkholderia cepaciawith Sustained Persistence in the Spleen
Authors: Speert David P. , Steen Barbara , Halsey Keith , Kwan Eddie
Published: December 2019

Burkholderia cepaciais an opportunistic pathogen that causes severe systemic infections in patients with chronic granulomatous disease (CGD) or with cystic fibrosis (CF), but its mechanisms of virulence are poorly understood. We developed a murine model of systemic infection in wild-type (WT) and gamma interferon knockout (GKO) BALB/c mice to facilitate dissection of components of pathogenicity and host defense. Both WT and GKO mice were susceptible to chronic splenic infection withB. cepacia, but not withPseudomonas aeruginosa. B. cepaciastrains from patients with CGD persisted longer than those from CF patients. C57BL/6 mice were the most susceptible murine strain; bacteria persisted in the spleen for 2 months. DBA/2, BALB/c, and A/J strains of mice were relatively resistant to infection. Certain strains ofB. cepaciacomplex can persist in the murine spleen after systemic infection; this may provide clues to its virulence in compromised hosts, such as those with CGD and CF.
TheTreponema denticolaMajor Sheath Protein Is Predominantly Periplasmic and Has Only Limited Surface Exposure
Authors: Caimano Melissa J. , Bourell Kenneth W. , Bannister Teresa D. , Cox David L. , Radolf Justin D.
Published: December 2019

The recent discovery that theTreponema pallidumgenome encodes 12 orthologs of theTreponema denticolamajor sheath protein (Msp) prompted us to reexamine the cellular location and topology of theT. denticolapolypeptide. Experiments initially were conducted to ascertain whether Msp forms an array on or within theT. denticolaouter membrane. Transmission electron microscopy (EM) of negatively stained and ultrathin-sectioned organisms failed to identify a typical surface layer, whereas freeze-fracture EM revealed that theT. denticolaouter membrane contains heterogeneous transmembrane proteins but no array. In contrast, a lattice-like structure was observed in vesicles released from mildly sonicated treponemes; combined EM and biochemical analyses demonstrated that this structure was the peptidoglycan sacculus. Immunoelectron microscopy (IEM) subsequently was performed to localize Msp inT. denticola. Examination of negatively stained whole mounts identified substantial amounts of Msp in sonicated organisms. IEM of ultrathin-sectioned, intact treponemes also demonstrated that the preponderance of antigen was unassociated with the outer membrane. Lastly, immunofluorescence analysis of treponemes embedded in agarose gel microdroplets revealed that only minor portions of Msp are surface exposed. Taken as a whole, our findings challenge the widely held belief that Msp forms an array within theT. denticolaouter membrane and demonstrate, instead, that it is predominantly periplasmic with only limited surface exposure. These findings also have implications for our evolving understanding of the contribution(s) of Msp/Tpr orthologs to treponemal physiology and disease pathogenesis.
Differential Regulation ofSalmonella typhimuriumType III Secreted Proteins by Pathogenicity Island 1 (SPI-1)-Encoded Transcriptional Activators InvF and HilA
Authors: Eichelberg Katrin , Galán Jorge E.
Published: December 2019

Salmonella entericaencodes a type III protein secretion system within a pathogenicity island (SPI-1) that is located at centisome 63 of its chromosome. This system is required for the ability of these bacteria to stimulate cellular responses that are essential for their pathogenicity. Expression of components and substrates of this system is subject to complex regulatory mechanisms. These mechanisms involve the function of HilA and InvF, two transcriptional regulatory proteins encoded within SPI-1. In this study, we examined the functional relationship between these two regulatory proteins. We found that strains carrying loss-of-function mutations in eitherhilAorinvFdiffer in their ability to stimulate cellular responses. AnS. typhimurium hilAmutant strain retained considerable signaling capacity that resulted in significant levels of internalization into host cells. In contrast, introduction of a nonpolar loss-of-function mutation ininvFrenderedS. typhimuriumsignificantly impaired in its ability to enter host cells. Consistent with these different phenotypes, we found that HilA and InvF control the expression of different genes. HilA regulates the expression of components of the type III secretion machinery, whereas InvF controls the expression of type III secreted proteins encoded outside of SPI-1. We also found that the expression of secreted proteins encoded within SPI-1 are under the control of both HilA and InvF. Our results therefore indicate that InvF and HilA differentially control the expression of components and substrates of the invasion-associated type III secretion system.
Intranasal Immunization with Pneumococcal Polysaccharide Conjugate Vaccines Protects Mice against Invasive Pneumococcal Infections
Authors: Jakobsen Håvard , Saeland Eiríkur , Gizurarson Sveinbjörn , Schulz Dominique , Jónsdóttir Ingileif
Published: December 2019

Host defenses against
Streptococcus pneumoniae
depend largely on opsonophagocytosis mediated by antibodies and complement. Since pneumococcus is a respiratory pathogen, mucosal immune responses may play a significant role in the defense against pneumococcal infections. Thus, mucosal vaccination may be an alternative approach to current immunization strategies, but effective adjuvants are required. Protein antigens induce significant mucosal immunoglobulin A (IgA) and systemic IgG responses when administered intranasally (i.n.) with the glyceride-polysorbate based adjuvant RhinoVax (RV) both in experimental animals and humans. The immunogenicity and efficacy of pneumococcal polysaccharide conjugate vaccines (PNC) of serotypes 1 and 3 was studied in mice after i.n. immunization with RV. Antibodies were measured in serum (IgM, IgG, and IgA) and saliva (IgA) and compared to antibody titers induced by parenteral immunization. The PNCs induced significant systemic IgG and IgA antibodies after i.n. immunization only when given with RV and, for serotype 1, serum IgG titers were comparable to titers induced by subcutaneous immunization. In addition, i.n. immunization with PNC-1 in RV elicited detectable mucosal IgA. These results demonstrate that RV is a potent mucosal adjuvant for polysaccharides conjugated to proteins. A majority of the PNC-1-immunized mice were protected against both bacteremia and pneumonia after i.n. challenge with a lethal dose of serotype 1 pneumococci, and protection correlated significantly with the serum IgG titers. Similarly, the survival of mice immunized i.n. with PNC-3 in RV was significantly prolonged. These results indicate that mucosal vaccination with PNC and adjuvants may be an alternative strategy for prevention against pneumococcal infections.
Elevated Chemokine Concentrations in Sera of Human Immunodeficiency Virus (HIV)-Seropositive and HIV-Seronegative Patients with Tuberculosis: a Possible Role for Mycobacterial Lipoarabinomannan
Authors: Juffermans Nicole P. , Verbon Annelies , van Deventer Sander J. H. , van Deutekom Henk , Belisle John T. , Ellis Michael E. , Speelman Peter , van der Poll Tom
Published: December 2019

Levels of interleukin 8 (IL-8), gamma interferon-inducible protein 10 (IP-10), monocyte chemoattractant protein 1 (MCP-1), and macrophage inflammatory protein 1β (MIP-1β) were elevated in patients with tuberculosis. IP-10 and MCP-1 levels were higher in human immunodeficiency virus (HIV)-seropositive patients than in HIV-seronegative patients with tuberculosis. Lipoarabinomannan induced IL-8, MCP-1, and MIP-1β in vitro, which was partly inhibited by anti-tumor necrosis factor antibody.
Iron Modulates Phenotypic Variation and Phosphorylation of P270 in Double-Stranded RNA Virus-Infected
Trichomonas vaginalis
Authors: Alderete J. F.
Published: December 2019

Trichomonas vaginalis
infected with a double-stranded RNA virus undergoes phenotypic variation on the basis of surface versus cytoplasmic expression of the immunogenic protein P270. Examination of batch cultures by flow cytofluorometry with monoclonal antibody (MAb) to P270 yields both fluorescent and nonfluorescent trichomonads. Greater numbers and intensity of fluorescent organisms with surface P270 reactive with MAb were evident in parasites grown in medium depleted of iron. Placement of iron-limited organisms in medium supplemented with iron gave increased numbers of nonfluorescent trichomonads. Purified subpopulations of trichomonads with and without surface P270 obtained by fluorescence-activated cell sorting reverted to nonfluorescent and fluorescent phenotypes when placed in high- and low-iron media, respectively. No similar regulation by iron of P270 was evident among virus-negative
T. vaginalis
isolates or virus-negative progeny trichomonads derived from virus-infected isolates. Equal amounts of P270 were detectable by MAb on immunoblots of total proteins from identical numbers of parasites grown in low- and high-iron media. Finally, P270 was found to be highly phosphorylated in high-iron parasites. Iron, therefore, plays a role in modulating surface localization of P270 in virus-harboring parasites.
Intranasal Administration of Synthetic Recombinant Peptide-Based Vaccine Protects Mice from Infection bySchistosoma mansoni
Authors: Ben-Yedidia T. , Tarrab-Hazdai R. , Schechtman D. , Arnon R.
Published: December 2019

Schistosomiasis is the cause of a chronic debilitating disease which accounts for significant mortality and morbidity every year, especially in tropical and subtropical areas. An epitope derived from the protective surface protein 9B-Ag ofSchistosoma mansoni, designated 9B peptide-1, was previously showed to be protective in mice when conjugated to bovine serum albumin and administered subcutaneously in complete Freund’s adjuvant. In this work, this protective peptide was expressed in the flagellin of aSalmonellavaccine strain, and the isolated recombinant flagella were used for immunization of mice. Since during the invasion of the parasite into the host the schistosomula migrate first to the lungs, the intranasal route of administration was employed in order to halt the parasite at an early stage of the infection. Such intranasal immunization with this peptide expressed in flagellin, without the addition of adjuvants, resulted in a significant humoral response and also led to protection against challenge infection, manifested as a reduction of the worm burden by an average of 42%.
A Defect in Interleukin-10 Leads to Enhanced Malarial Disease inPlasmodium chabaudi chabaudiInfection in Mice
Authors: Li Ching , Corraliza Inés , Langhorne Jean
Published: December 2019

Infection of interleukin-10 (IL-10)-nonexpressing (IL-10−/−) mice withPlasmodium chabaudi chabaudi(AS) leads to exacerbated pathology in female mice and death in a proportion of them. Hypoglycemia, hypothermia, and loss in body weight were significantly greater in female IL-10−/−mice than in male knockout mice and all wild-type (WT) mice during the acute phase of infection. At this time, both female and male IL-10−/−mice produced more gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), and IL-12p40 mRNA than their respective WT counterparts. Inactivation of IFN-γ in IL-10−/−mice by the injection of anti-IFN-γ antibodies or by the generation of IL-10−/−IFN-γ receptor−/−double-knockout mice resulted in reduced mortality but did not affect body weight, temperature, or blood glucose levels. The data suggest that IFN-γ-independent pathways may be responsible for these pathological features ofP. chabaudimalaria and may be due to direct stimulation of TNF-α by the parasite. Since male and female knockout mice both produce more inflammatory cytokines than their WT counterparts, it is likely that the mortality seen in females is due to the nature or magnitude of the response to these cytokines rather than the amount of IFN-γ or TNF-α produced.
Reactivation of Latent Tuberculosis: Variations on the Cornell Murine Model
Authors: Scanga Charles A. , Mohan V. P. , Joseph Heather , Yu Keming , Chan John , Flynn JoAnne L.
Published: December 2019

Mycobacterium tuberculosiscauses active tuberculosis in only a small percentage of infected persons. In most cases, the infection is clinically latent, although immunosuppression can cause reactivation of a latentM. tuberculosisinfection. Surprisingly little is known about the biology of the bacterium or the host during latency, and experimental studies on latent tuberculosis suffer from a lack of appropriate animal models. The Cornell model is a historical murine model of latent tuberculosis, in which mice infected withM. tuberculosisare treated with antibiotics (isoniazid and pyrazinamide), resulting in no detectable bacilli by organ culture. Reactivation of infection during this culture-negative state occurred spontaneously and following immunosuppression. In the present study, three variants of the Cornell model were evaluated for their utility in studies of latent and reactivated tuberculosis. The antibiotic regimen, inoculating dose, and antibiotic-free rest period prior to immunosuppression were varied. A variety of immunosuppressive agents, based on immunologic factors known to be important to control of acute infection, were used in attempts to reactivate the infection. Although reactivation of latent infection was observed in all three variants, these models were associated with characteristics that limit their experimental utility, including spontaneous reactivation, difficulties in inducing reactivation, and the generation of altered bacilli. The results from these studies demonstrate that the outcome of Cornell model-based studies depends critically upon the parameters used to establish the model.
Characterization of VspB of
Borrelia turicatae
, a Major Outer Membrane Protein Expressed in Blood and Tissues of Mice
Authors: Pennington Pamela M. , Cadavid Diego , Barbour Alan G.
Published: December 2019

Serotypes A and B of the relapsing fever spirochete
Borrelia turicatae
produce different disease manifestations in infected mice. Whereas serotype B causes more severe arthritis and reaches higher densities in the blood of mice than serotype A, serotype A invades the central nervous system earlier than serotype B during infection. These differences between serotypes A and B in mice are associated with the expression of different surface proteins, VspA and VspB, respectively, in the culture medium. To determine whether these proteins, in particular, VspB, are also expressed in vivo,
scid
mice infected with
B. turicatae
were studied. The expression of VspB by spirochetes in the blood was demonstrated in Coomassie blue-stained polyacrylamide gels and Western blots with a specific monoclonal antibody. Indirect immunofluorescence and immunoperoxidase studies confirmed the expression of VspB in the blood and also demonstrated VspB expression in the joints and heart. The gene for VspB was next identified and cloned by using partial amino acid sequencing, reverse transcriptase PCR, and a specific monoclonal antibody. The
vspB
gene encodes a protein of 216 amino acids that is 68% identical to VspA of
B. turicatae
and 44 to 56% identical to representative Vsp and OspC lipoproteins of other
Borrelia
spp. The processed VspB protein was distinguished from 26 other Vsp and OspC proteins by a high predicted isoelectric point at 9.39. The promoter region for
vspB
was similar to the promoter region for the
vsp33
gene of
Borrelia hermsii
and for the
ospC
gene of
Borrelia burgdorferi
, two genes known to be environmentally regulated. These studies established that the virulence-associated VspB protein is expressed by spirochetes in the mouse and that VspB is a novel member of the Vsp-OspC family of proteins.
Control of White-Opaque Phenotypic Switching in
Candida albicans
by the Efg1p Morphogenetic Regulator
Authors: Sonneborn Anja , Tebarth Bernd , Ernst Joachim F.
Published: December 2019

Phenotypic switching in
Candida albicans
spontaneously generates different cellular morphologies and is manifested in strain WO-1 by the reversible switching between the white and opaque phenotypes. We present evidence that phenotypic switching is regulated by the Efg1 protein, which is known as an essential element of hyphal development (dimorphism). Firstly,
EFG1
is expressed specifically in cells of the white but not the opaque phenotype. During mass conversion from the opaque to the white phenotype, the
EFG1
transcript level correlates with competence of switching of opaque cells to the white form. Secondly, overexpression of
EFG1
by a
PCK1p-EFG1
fusion forces opaque-phase cells to switch to the white form with a high level of efficiency. Thirdly, low-level expression of
EFG1
in strain CAI-8 generates a cellular phenotype similar to that of opaque cells in that cells bud as short rods, which cannot be induced to form hyphae in standard conditions; such cells (unlike authentic opaque cells) lack typical surface “pimples.” Importantly, the opaque-specific
OP4
transcript is induced in the opaque-like cells generated by strain CAI8 as a response to low-level expression of
EFG1
. The results suggest that high
EFG1
expression levels induce and maintain the white cell form while low
EFG1
expression levels induce and maintain the opaque cell form. It is proposed that changes in
EFG1
expression determine or contribute to phenotypic switching events in
C. albicans
.
Genetic Control of Schistosome Infections by the SM1 Locus of the 5q31-q33 Region Is Linked to Differentiation of Type 2 Helper T Lymphocytes
Authors: Rodrigues Virmondes , Piper Karen , Couissinier-Paris Patricia , Bacelar Olivia , Dessein Hélia , Dessein Alain J.
Published: December 2019

Human susceptibility toSchistosoma mansoniinfections is controlled by the SM1 locus on chromosome 5 in q31-q33. This genetic region encodes cytokines which regulate the development of helper T lymphocytes. In the present work, a clonal analysis of CD4+T lymphocytes of homozygous resistant and homozygous susceptible subjects was undertaken to evaluate whether SM1 controls helper T-cell differentiation. Of 121 CD4+T-cell clones (TCC) from three susceptible (S) and three resistant (R) subjects, 68 proliferated when stimulated by parasite antigens. Parasite-specific TCC derived from susceptible subjects (33 STCC) produced 10- to 1,000-fold less interleukin-4 and -5 than TCC from resistant subjects (25 RTCC). Clones from both patient groups produced, however, the same amount of gamma interferon. Parasite-specific STCC were type 1 helper (Th1) or Th0/1, whereas RTCC were either Th2 or Th0/2. These results, together with the localization of SM1 in 5q31-q33, indicate that the SM1 locus controls the differentiation of Th2 lymphocytes.
Identification, Cloning, and Expression of the CAMP factor gene (
cfa
) of Group A Streptococci
Authors: Gase Klaus , Ferretti Joseph J. , Primeaux Charles , McShan W. Michael
Published: December 2019

The CAMP reaction is a synergistic lysis of erythrocytes by the interaction of an extracellular protein (CAMP factor) produced by some streptococcal species with the
Staphylococcus aureus
sphingomyelinase C (beta-toxin). Group A streptococci (GAS [
Streptococcus pyogenes
]) have been long considered CAMP negative, and this reaction commonly has been used to distinguish GAS from
Streptococcus agalactiae
. We here provide evidence that GAS possess this gene and produce an extracellular CAMP factor capable of participating in a positive CAMP reaction. The
S. pyogenes
CAMP factor is specified by a 774-bp open reading frame homologous to the CAMP factor genes from
S. agalactiae
and
Streptococcus uberis
. This gene, designated
cfa
, was isolated on a 1,256-bp fragment and cloned in
Escherichia coli
. Recombinant clones of
E. coli
expressing
cfa
secreted an active CAMP factor. The deduced 28.5-kDa protein encoded by
cfa
consists of 257 amino acids, with a predicted 28-amino-acid signal peptide. The
cfa
gene is widely spread among GAS: 82 of 100 clinical GAS isolates produced a positive CAMP reaction. Of the CAMP-negative strains, 17 of the 18 GAS strains contained the
cfa
gene. Additionally, CAMP activity was detected in streptococci from serogroups C, M, P, R, and U. The
cfa
gene was cloned and actively expressed in
Escherichia coli
and gene fusions were made, placing the β-galactosidase gene (
lacZ
) under control of the
cfa
promoter. These
cfa
promoter-
lacZ
fusions were introduced into
S. pyogenes
via a bacteriophage-derived site-specific integration vector where they showed that the
cfa
gene has a strong promoter that may be subject to as-yet-unidentified regulatory factors. The results presented here, along with previous reports, indicate that the CAMP factor gene is fairly widespread among streptococci, being present at least in groups A, B, C, G, M, P, R, and U.
T-Cell Responses toTreponema pallidumsubsp.pallidumAntigens during the Course of Experimental Syphilis Infection
Authors: Arroll Thomas W. , Centurion-Lara Arturo , Lukehart Sheila A. , Van Voorhis Wesley C.
Published: December 2019

In this study we describe the development of the T-cell response to a panel ofTreponema pallidumantigens over the course of syphilis infection in the rabbit and determine whether these antigens induce the expression of Th1 cytokines. It was determined that the membrane proteins TpN17 and TpN47, as well as the endoflagellar sheath protein TpN37, induce strong proliferation responses through most of syphilis infection; Tromp1 induced only weak proliferative responses. An unexpected drop in proliferative response to these antigens at day 90 of infection, followed by a dramatic increase in response at day 180, suggests that there may be a secondary dissemination ofT. pallidumwhich induces a recall response. Crude epitope mapping of TpN17 and TpN37 showed that multiple epitopes may be present on both antigens, which is likely a contributing factor in the immunodominance of these antigens. The T-cell response to the TpN37 molecule shows acquisition of newly recognized epitopes during the course of infection. SonicatedT. pallidumwas found to induce the expression of interleukin-2 (IL-2) and gamma interferon and not IL-10 mRNA, showing that the general T-cell response toT. pallidumantigens in syphilis infection is biased towards the Th1 phenotype. Of the antigens tested, TpN37 appears to contribute the most to the Th1 cytokine response and therefore may play a key role in the clearance ofT. pallidumfrom lesions.
LcrV ofYersinia pestisEnters Infected Eukaryotic Cells by a Virulence Plasmid-Independent Mechanism
Authors: Fields Kenneth A. , Straley Susan C.
Published: December 2019

Yersinia pestisis the causative agent of bubonic plague and possesses a set of plasmid-encoded, secretable virulence proteins termed LcrV and Yops which are essential for survival in mammalian hosts. Yops and LcrV are secreted by a type III mechanism (Ysc), and Yops are unidirectionally targeted into the cytosol of associated eukaryotic cells in a tissue culture infection model. LcrV is required for Yops targeting, and recent findings have revealed that it can localize to the bacterial surface; however, its fate in this infection model has not been investigated in detail. In this study, we compared the localization of LcrV to that of the targeted proteins YopE and YopM by immunoblot analysis of fractions ofYersinia-infected HeLa cultures or by laser-scanning confocal microscopy of infected monolayers. Both LcrV and YopE were secreted by contact-activated, extracellularly localized yersiniae and were targeted to the HeLa cell cytosol. Although a significant amount of LcrV partitioned to the culture medium (unlike YopE), this extracellular pool of LcrV was not the source of the LcrV that entered HeLa cells. Unlike targeting of YopE and YopM, targeting of LcrV occurred in the absence of a functional Ysc apparatus and other virulence plasmid (pCD1)-expressed proteins. However, the Ysc is necessary for LcrV to be released into the medium, and our recent work has shown that localization of LcrV on the bacterial surface requires the Ysc. These results indicate that two mechanisms exist for the secretion of LcrV byY. pestis, both of which are activated by contact with eukaryotic cells. LcrV secreted by the Ysc reaches the bacterial surface and the surrounding medium, whereas the second is a novel, Ysc-independent pathway which results in localization of LcrV in the cytosol of infected cells but not the surrounding medium.
Contribution ofSalmonella typhimuriumVirulence Factors to Diarrheal Disease in Calves
Authors: Tsolis Renée M. , Adams L. Garry , Ficht Thomas A. , Bäumler Andreas J.
Published: December 2019

Limited knowledge is available about the virulence mechanisms responsible for diarrheal disease caused bySalmonella typhimurium. To assess the contribution to diarrheal disease of virulence determinants identified in models of infection, we tested a collection ofS. typhimuriummutants for their ability to cause enteritis in calves.S. typhimuriumstrains carrying mutations in the virulence plasmid (spvR),Salmonellapathogenicity island 2 (SPI-2) (spiB), or SPI-5 (sopB) caused mortality and acute diarrhea in calves. AnS. typhimurium rfaJmutant, which is defective for lipopolysaccharide outer core biosynthesis, was of intermediate virulence. Mutations in SPI-1 (hilAandprgH) oraroAmarkedly reduced virulence and the severity of diarrhea. Furthermore, histopathological examination of calves infected with SPI-1 oraroAmutants revealed a marked reduction or absence of intestinal lesions. These data suggest that virulence factors, such as SPI-1, which are required during intestinal colonization are more important for pathogenicity in calves than are genes required during the systemic phase ofS. typhimuriuminfection, including SPI-2 or thespvoperon. This is in contrast to the degree of attenuation caused by these mutations in the mouse.
Use of Genetically Manipulated Strains of
Clostridium perfringens
Reveals that Both Alpha-Toxin and Theta-Toxin Are Required for Vascular Leukostasis To Occur in Experimental Gas Gangrene
Authors: Ellemor Darren M. , Baird Rebecca N. , Awad Milena M. , Boyd Richard L. , Rood Julian I. , Emmins John J.
Published: December 2019

A hallmark of gas gangrene (clostridial myonecrosis) pathology is a paucity of leukocytes infiltrating the necrotic tissue. The cause of this paucity most likely relates to the observation of leukocyte aggregates at the border of the area of tissue necrosis, often within the microvasculature itself. Infecting mice with genetically manipulated strains of
Clostridium perfringens
type A (deficient in either alpha-toxin or theta-toxin production) resulted in significantly reduced leukocyte aggregation when alpha-toxin was absent and complete abrogation of leukocyte aggregation when theta-toxin was absent. Thus, both alpha-toxin and theta-toxin are necessary for the characteristic vascular leukostasis observed in clostridial myonecrosis.
Role of Local Cytokines in Increased Gastric Expression of the Secretory Component inHelicobacter pyloriInfection
Authors: Ahlstedt Ingela , Lindholm Catharina , Lönroth Hans , Hamlet Annika , Svennerholm Ann-Mari , Quiding-Järbrink Marianne
Published: December 2019

Using immunohistochemical staining, we examined the presence of secretory component (SC) on epithelial cells in gastric and duodenal biopsy specimens collected fromHelicobacter pylori-infected individuals and healthy controls. Gastric epithelial cells from healthy volunteers expressed low, but detectable, levels of SC. In contrast, significantly higher level of expression of SC (P< 0.001) was observed on epithelial cells in the antra ofH. pylori-infected individuals. The antral SC expression correlated with staining for gamma interferon of intraepithelial and lamina propria lymphocytes (rs= 0.76 and 0.69, respectively,P< 0.001) and correlated weakly with production of tumor necrosis factor alpha (rs= 0.43,P< 0.05), but it did not correlate at all with interleukin-4 production.
Age-Associated Differences in Immunoglobulin G1 (IgG1) and IgG2 Subclass Antibodies to Pneumococcal Polysaccharides following Vaccination
Authors: Lottenbach Kathleen R. , Mink ChrisAnna M. , Barenkamp Stephen J. , Anderson Edwin L. , Homan Sharon M. , Powers Douglas C.
Published: December 2019

Immunoglobulin G (IgG) subclass antibody responses to pneumococcal vaccines were determined for human subjects in four age groups. The ratios of IgG1/IgG2 antibody concentrations declined with advancing age for all five of the serotypes tested. Protein-conjugate vaccines elicited enhanced IgG antibody responses over plain polysaccharide vaccines in infants but not in adult groups.
Coinfection withToxoplasma gondiiInhibits Antigen-Specific Th2 Immune Responses, Tissue Inflammation, and Parasitism in BALB/c Mice Infected withLeishmania major
Authors: Santiago H. C. , Oliveira M. A. P. , Bambirra E. A. , Faria A. M. C. , Afonso L. C. C. , Vieira L. Q. , Gazzinelli R. T.
Published: December 2019

Lesion size, cellular infiltration, and tissue parasitism in the footpads of BALB/c mice infected withLeishmania majorwere all dramatically inhibited during acute but not chronic infection withToxoplasma gondii. Similarly, acute but not chronic toxoplasmosis at the time of infection withL. majorhad a strong inhibitory effect on development of acquired immune responses mediated by Th2 lymphocytes. In contrast, no major changes inLeishmania-specific Th1-mediated responses were observed in mice coinfected withT. gondii.
Identification of a Third Metalloprotease Toxin Gene in Extraintestinal Isolates of
Bacteroides fragilis
Authors: Chung Gyung-Tae , Franco Augusto A. , Wu Shaoguang , Rhie Gi-Eun , Cheng Rodney , Oh Hee-Bok , Sears Cynthia L.
Published: December 2019

To further understand the epidemiology of enterotoxigenic
Bacteroides fragilis
(ETBF), 89 extraintestinal
B. fragilis
strains from Seoul, Korea, were examined for secretion of
B. fragilis
toxin (BFT) by the HT29/C1 biologic assay and for the
B. fragilis
toxin gene (
bft
) by colony blot hybridization and PCR. Complete agreement between the three techniques was found. Overall, 34
B. fragilis
strains (38%) were identified as ETBF. Eleven of the 34 ETBF strains (32%) expressed a new isoform of BFT (Korea-BFT). This new isoform is more related to BFT-2 than to BFT-1. Like BFT-1 and BFT-2, Korea-BFT cleaves E-cadherin, the zonula adherens protein.
Immune Cells Are Required for Cutaneous Ulceration in a Swine Model of Chancroid
Authors: San Mateo Lani R. , Toffer Kristen L. , Orndorff Paul E. , Kawula Thomas H.
Published: December 2019

Cutaneous lesions of the human sexually transmitted genital ulcer disease chancroid are characterized by the presence of intraepidermal pustules, keratinocyte cytopathology, and epidermal and dermal erosion. These lesions are replete with neutrophils, macrophages, and CD4
+
T cells and contain very low numbers of cells of
Haemophilus ducreyi
, the bacterial agent of chancroid. We examined lesion formation by
H. ducreyi
in a pig model by using cyclophosphamide (CPA)-induced immune cell deficiency to distinguish between host and bacterial contributions to chancroid ulcer formation. Histologic presentation of
H. ducreyi
-induced lesions in CPA-treated pigs differed from ulcers that developed in immune-competent animals in that pustules did not form and surface epithelia remained intact. However, these lesions had significant suprabasal keratinocyte cytotoxicity. These results demonstrate that the host immune response was required for chancroid ulceration, while bacterial products were at least partially responsible for the keratinocyte cytopathology associated with chancroid lesions in the pig. The low numbers of
H. ducreyi
present in lesions in humans and immune-competent pigs have prevented localization of these organisms within skin. However,
H. ducreyi
organisms were readily visualized in lesion biopsies from infected CPA-treated pigs by immunoelectron microscopy. These bacteria were extracellular and associated with necrotic host cells in the epidermis and dermis. The relative abundance of
H. ducreyi
in inoculated CPA-treated pig skin suggests control of bacterial replication by host immune cells during natural human infection.
Glycerol Monolaurate Inhibits Induction of Vancomycin Resistance in
Enterococcus faecalis
Authors: Ruzin Alexey , Novick Richard P.
Published: December 2019

Glycerol monolaurate (GML) is a surfactant that has been found to inhibit the post-exponential phase activation of virulence factor production and the induction of β-lactamase in
Staphylococcus aureus
. It has been suggested that signal transduction is the most probable target for GML (S. J. Projan, S. Brown-Skrobot, P. M. Schlievert, F. Vandenesch, and R. P. Novick, J. Bacteriol. 176:4204–4209, 1994). We found that GML suppresses growth of vancomycin-resistant
Enterococcus faecalis
on plates with vancomycin and blocks the induction of vancomycin resistance, which involves a membrane-associated signal transduction mechanism, either at or before initiation of transcription. Given the surfactant nature of GML and the results of previous experiments, we suggest that GML blocks signal transduction. In contrast, GML has no effect on the induction of erythromycin-inducible macrolide resistance in
S. aureus
, which does not involve signal transduction.
The
leuO
Gene Product Has a Latent Ability To Relieve
bgl
Silencing in
Escherichia coli
Authors: Ueguchi Chiharu , Ohta Tomoko , Seto Chiharu , Suzuki Tomomi , Mizuno Takeshi
Published: December 2019

The
Escherichia coli bgl
operon is of interest, since its expression is silent (phenotypically Bgl
−
), at least under standard laboratory conditions. Here we attempted to identify a
trans
-acting factor(s) that is presumably relevant to the regulation of
bgl
by a random insertion mutagenesis with mini-Tn
10
. These collected mutations, conferring the phenotype of Bgl
+
, were localized in three loci on the genetic map, two of which appeared to be
hns
and
bglJ
, which were previously implicated as the factors affecting the Bgl phenotype. The other locus at 1 to 2 min on the genetic map appeared to be a new one. In this case, the insertion mutation was found to be just in front of the
leuO
gene encoding a putative LysR-like DNA-binding protein. Genetic analyses revealed that overproduction of LeuO in the wild-type cells causes the phenotype of Bgl
+
. A
leuO
deletion mutant was also characterized in terms of expression of
bgl
. From these results, the possible function of LeuO in
bgl
expression will be discussed from an evolutionary and/or ecological point of view.
EpsR Modulates Production of Extracellular Polysaccharides in the Bacterial Wilt Pathogen
Ralstonia
(
Pseudomonas
)
solanacearum
Authors: Chapman Matt R. , Kao C. Cheng
Published: December 2019

Ralstonia solanacearum
is the causal agent of bacterial wilt of many agriculturally important crops. Exopolysaccharide synthesized by products of the
epsI
operon is the major virulence factor for
R. solanacearum
. Expression of
epsI
has been demonstrated to be under the control of several proteins, including several two-component regulators. Overexpression of EpsR was found previously to reduce the amount of synthesis specifically from the
epsI
promoter. Here we present data that a single chromosomal copy of
epsR
activates the
epsI
promoter, suggesting that EpsR is a concentration-dependent effector of
epsI
gene expression. Furthermore, the ability of EpsR to modulate
epsI
expression is dependent on the phosphorylation state of EpsR. Gel mobility shift assays suggest that EpsR can specifically bind the
epsI
promoter and that this binding requires a phosphorylated form of EpsR.