bluuebunny/crossref_metadata_2025_split huggingface dataset:
Eradication of Intracellular
Salmonella
enterica
Serovar Typhimurium with a Small-Molecule, Host Cell-Directed Agent
Authors: Chiu Hao-Chieh , Kulp Samuel K. , Soni Shilpa , Wang Dasheng , Gunn John S. , Schlesinger Larry S. , Chen Ching-Shih 
Published: October 2009

ABSTRACT

Eradication of intracellular pathogenic bacteria with host-directed chemical agents has been an anticipated innovation in the treatment of antibiotic-resistant bacteria. We previously synthesized and characterized a novel small-molecule agent, AR-12, that induces autophagy and inhibits the Akt kinase in cancer cells. As both autophagy and the Akt kinase have been shown recently to play roles in the intracellular survival of several intracellular bacteria, including
Salmonella
enterica
serovar Typhimurium, we investigated the effect of AR-12 on the intracellular survival of
S
almonella
serovar Typhimurium in macrophages. Our results show that AR-12 induces autophagy in macrophages, as indicated by increased autophagosome formation, and potently inhibits the survival of serovar Typhimurium in macrophages in association with increased colocalization of intracellular bacteria with autophagosomes. Intracellular bacterial growth was partially rescued in the presence of AR-12 by the short hairpin RNA-mediated knockdown of Beclin-1 or Atg7 in macrophages. Moreover, AR-12 inhibits Akt kinase activity in infected macrophages, which we show to be important for its antibacterial effect as the enforced expression of constitutively activated Akt1 in these cells reverses the AR-12-induced inhibition of intracellular serovar Typhimurium survival. Finally, oral administration of AR-12 at 2.5 mg/kg/day to serovar Typhimurium-infected mice reduced hepatic and splenic bacterial burdens and significantly prolonged survival. These findings show that AR-12 represents a proof of principle that the survival of intracellular bacteria can be suppressed by small-molecule agents that target both innate immunity and host cell factors modulated by bacteria.
In Vitro Activity of Gallium Maltolate against Staphylococci in Logarithmic, Stationary, and Biofilm Growth Phases: Comparison of Conventional and Calorimetric Susceptibility Testing Methods
Authors: Baldoni Daniela , Steinhuber Andrea , Zimmerli Werner , Trampuz Andrej 
Published: October 2009

ABSTRACTGa3+is a semimetal element that competes for the iron-binding sites of transporters and enzymes. We investigated the activity of gallium maltolate (GaM), an organic gallium salt with high solubility, against laboratory and clinical strains of methicillin-susceptibleStaphylococcus aureus(MSSA), methicillin-resistantS. aureus(MRSA), methicillin-susceptibleStaphylococcus epidermidis(MSSE), and methicillin-resistantS. epidermidis(MRSE) in logarithmic or stationary phase and in biofilms. The MICs of GaM were higher forS. aureus(375 to 2000 μg/ml) thanS. epidermidis(94 to 200 μg/ml). Minimal biofilm inhibitory concentrations were 3,000 to ≥6,000 μg/ml (S. aureus) and 94 to 3,000 μg/ml (S. epidermidis). In time-kill studies, GaM exhibited a slow and dose-dependent killing, with maximal action at 24 h againstS. aureusof 1.9 log10CFU/ml (MSSA) and 3.3 log10CFU/ml (MRSA) at 3× MIC and 2.9 log10CFU/ml (MSSE) and 4.0 log10CFU/ml (MRSE) againstS. epidermidisat 10× MIC. In calorimetric studies, growth-related heat production was inhibited by GaM at subinhibitory concentrations; and the minimal heat inhibition concentrations were 188 to 4,500 μg/ml (MSSA), 94 to 1,500 μg/ml (MRSA), and 94 to 375 μg/ml (MSSE and MRSE), which correlated well with the MICs. Thus, calorimetry was a fast, accurate, and simple method useful for investigation of antimicrobial activity at subinhibitory concentrations. In conclusion, GaM exhibited activity against staphylococci in different growth phases, including in stationary phase and biofilms, but high concentrations were required. These data support the potential topical use of GaM, including its use for the treatment of wound infections, MRSA decolonization, and coating of implants.
Transcriptomic Profiling of theSaccharomyces cerevisiaeResponse to Quinine Reveals a Glucose Limitation Response Attributable to Drug-Induced Inhibition of Glucose Uptake
Authors: dos Santos Sandra C. , Tenreiro Sandra , Palma Margarida , Becker Jorg , Sá-Correia Isabel 
Published: October 2009

ABSTRACTQuinine has been employed in the treatment of malaria for centuries and is still used against severePlasmodium falciparummalaria. However, its interactions with the parasite remain poorly understood and subject to debate. In this study, we used theSaccharomyces cerevisiaeeukaryotic model to better understand quinine's mode of action and the mechanisms underlying the cell response to the drug. We obtained a transcriptomic profile of the yeast's early response to quinine, evidencing a marked activation of genes involved in the low-glucose response (e.g.,CAT8,ADR1,MAL33,MTH1, andSNF3). We used a low inhibitory quinine concentration with no detectable effect on plasma membrane function, consistent with the absence of a general nutrient starvation response and suggesting that quinine-induced glucose limitation is a specific response. We have further shown that transport of [14C]glucose is inhibited by quinine, with kinetic data indicating competitive inhibition. Also, tested mutant strains deleted for genes encoding high- and low-affinity hexose transporters (HXT1toHXT5,HXT8, andHXT10) exhibit resistance phenotypes, correlating with reduced levels of quinine accumulation in the mutants examined. These results suggest that the hexose transporters are facilitators of quinine uptake inS. cerevisiae, possibly through a competitive inhibition mechanism. Interestingly,P. falciparumis highly dependent on glucose uptake, which is mediated by the single-copy transporter PfHT1, a protein with high homology to yeast's hexose transporters. We propose that PfHT1 is an interesting candidate quinine target possibly involved in quinine import inP. falciparum, an uptake mechanism postulated in recent studies to occur through a still-unidentified importer(s).
Clindamycin-Induced CovS-Mediated Regulation of the Production of Virulent Exoproteins Streptolysin O, NAD Glycohydrolase, and Streptokinase in
Streptococcus pyogenes
Authors: Minami Masaaki , Kamimura Takuya , Isaka Masanori , Tatsuno Ichiro , Ohta Michio , Hasegawa Tadao 
Published: October 2009

ABSTRACT

The administration of high-dose clindamycin (CLI) along with penicillin is recommended for the treatment of streptococcal toxic shock syndrome. However, the prevalence of CLI-resistant
Streptococcus pyogenes
strains is increasing worldwide, and the effect of CLI on CLI-resistant
S. pyogenes
strains remains unknown. We aimed to evaluate the effect of CLI on the in vitro production of three major virulent exoproteins, namely, streptolysin O (Slo), NAD glycohydrolase (Nga), and streptokinase (Ska), by CLI-resistant
S. pyogenes
strains. After the incubation of M1 serotype CLI-resistant
S. pyogenes
D2TY in the presence of 1 μg/ml CLI, the amounts of Slo, Nga, and Ska and the levels of
slo
,
nga
, and
ska
mRNA in the supernatant were analyzed by Northern blotting and Western blotting, respectively. The results of both assays showed that the production of Slo, Nga, and Ska was higher with CLI treatment than without CLI treatment. We evaluated the role of the sensor kinase CovS, which is involved in the two-component system of
S. pyogenes
, in the CLI-induced production of these three exoproteins. Northern blotting analysis revealed that CLI induced the expression of
covS
mRNA in wild-type strain D2TY. Furthermore, both Northern blotting and Western blotting analyses showed that CLI decreased the levels of expression of Slo, Nga, and Ska in isogenic
covS
mutant D2TYcovS. These results suggest that CLI increases the production of three virulent exoproteins in CLI-resistant
S. pyogenes
strains via the action of CovS.
Mitochondrial DNA Depletion and Respiratory Chain Activity in Primary Human Subcutaneous Adipocytes Treated with Nucleoside Analogue Reverse Transcriptase Inhibitors
Authors: Stankov Metodi V. , Lücke Thomas , Das Anibh M. , Schmidt Reinhold E. , Behrens Georg M. N. 
Published: October 2009

ABSTRACTMitochondrial dysfunction as a consequence of mitochondrial DNA (mtDNA) depletion due to therapy with nucleoside analogue reverse transcriptase inhibitors (NRTI) has been proposed as a pathogenic mechanism leading to lipoatrophy in HIV-infected patients. The aim of our study was to investigate the impact of NRTI treatment on mtDNA abundance and the activities of respiratory chain complexes in primary human subcutaneous preadipocytes (phsPA). We studied adipocyte phenotypes, viability, and differentiation (CCAAT/enhancer-binding protein α [C/EBPα] and peroxisome proliferator-activated receptor γ [PPARγ] expression) and adiponectin production, mtDNA content, mitochondrial membrane potential, mitochondrial mass, and respiratory chain enzyme and citrate synthase activities in both proliferating and differentiating phsPA. Cells were exposed to zidovudine (6 μM), stavudine (d4T; 3 μM), and zalcitabine (ddC; 0.1 μM) for 8 weeks. NRTI-induced mtDNA depletion occurred in proliferating and differentiating phsPA after exposure to therapeutic drug concentrations of d4T and ddC. At these concentrations, ddC and d4T led to an almost 50% decrease in the number of mtDNA copies per cell without major impact on adipocyte differentiation. Despite mtDNA depletion by NRTI, the activities of the respiratory chain complexes, the mitochondrial membrane potential, and the mitochondrial mass were found to be unaffected. Severe NRTI-mediated mtDNA depletion in phsPA is not inevitably associated with impaired respiratory chain activity or altered mitochondrial membrane potential.
Prevention and Treatment ofStaphylococcus aureusPneumonia with a β-Cyclodextrin Derivative
Authors: Ragle Brook E. , Karginov Vladimir A. , Bubeck Wardenburg Juliane 
Published: October 2009

ABSTRACTStaphylococcus aureuspneumonia is a common, potentially life-threatening infection caused by this human pathogen. The only therapies available to treatS. aureuspneumonia are antibiotics, a modality that is jeopardized by the organism's remarkable ability to acquire antimicrobial resistance.S. aureusalpha-hemolysin is a pore-forming cytotoxin that is essential for the pathogenesis of pneumonia. Strains lacking this cytotoxin are avirulent in a murine model of pneumonia; likewise, vaccine-based strategies that antagonize the toxin afford protection against lethal disease. Disruption of the function of this toxin therefore provides a potent mechanism to prevent and/or treatS. aureuspneumonia. β-Cyclodextrin derivatives are small molecules with a sevenfold symmetry that mirrors the heptameric alpha-hemolysin. These compounds block the assembled alpha-hemolysin pore, compromising toxin function. We report that a modified β-cyclodextrin compound, IB201, prevents alpha-hemolysin-induced lysis of human alveolar epithelial cells. This protective effect does not result from the ability of the β-cyclodextrin to impair formation of the oligomeric alpha-hemolysin on the cell surface, supporting a role for this molecule in blockade of the lytic pore. An examination of IB201 in murineS. aureuspneumonia demonstrated that administration of this compound prevents and treats disease, protecting against mortality. Consistent with the vital importance of alpha-hemolysin in pneumonia caused by methicillin-sensitive and highly virulent methicillin-resistantS. aureusstrains, IB201 protects against lethal challenge with both types of isolates. These observations, coupled with a favorable safety profile of β-cyclodextrin compounds, provide a novel strategy that may be developed to combatS. aureuspneumonia.
Global Challenge of Antibiotic-Resistant
Treponema pallidum
Authors: Stamm Lola V. 
Published: October 2009

ABSTRACT

Syphilis is a multistage infectious disease that is usually transmitted through contact with active lesions of a sexual partner or from an infected pregnant woman to her fetus. Despite elimination efforts, syphilis remains endemic in many developing countries and has reemerged in several developed countries, including China, where a widespread epidemic recently occurred. In the absence of a vaccine, syphilis control is largely dependent upon identification of infected individuals and treatment of these individuals and their contacts with antibiotics. Although penicillin is still effective, clinically significant resistance to macrolides, a second-line alternative to penicillin, has emerged. Macrolide-resistant strains of
Treponema pallidum
are now prevalent in several developed countries. An understanding of the genetic basis of
T. pallidum
antibiotic resistance is essential to enable molecular surveillance. This review discusses the genetic basis of
T. pallidum
macrolide resistance and the potential of this spirochete to develop additional antibiotic resistance that could seriously compromise syphilis treatment and control.
The Sca2 Autotransporter Protein from
Rickettsia conorii
Is Sufficient To Mediate Adherence to and Invasion of Cultured Mammalian Cells
Authors: Cardwell Marissa M. , Martinez Juan J. 
Published: October 2009

ABSTRACT

Obligate intracellular bacteria of the genus
Rickettsia
must adhere to and invade the host endothelium in order to establish an infection. These processes require the interaction of rickettsial surface proteins with mammalian host cell receptors. A previous bioinformatic analysis of sequenced rickettsial species identified a family of at least 17 predicted “surface cell antigen” (
sca
) genes whose products resemble autotransporter proteins. Two members of this family, rOmpA and rOmpB of spotted fever group (SFG) rickettsiae have been identified as adhesion and invasion factors, respectively; however, little is known about the putative functions of the other
sca
gene products. An intact
sca2
gene is found in the majority of pathogenic SFG rickettsiae and, due to its sequence conservation among these species, we predict that Sca2 may play an important function at the rickettsial surface. Here we have shown that
sca2
is transcribed and expressed in
Rickettsia conorii
and have used a heterologous gain-of-function assay in
E. coli
to determine the putative role of Sca2. Using this system, we have demonstrated that expression of Sca2 at the outer membrane of nonadherent, noninvasive
E. coli
is sufficient to mediate adherence to and invasion of a panel of mammalian cells, including endothelial cells. Furthermore, soluble Sca2 protein is capable of diminishing
R. conorii
invasion of cultured mammalian cells. This is the first evidence that Sca2 participates in the interaction between SFG rickettsiae and host cells and suggests that in addition to other surface proteins, Sca2 may play a critical role in rickettsial pathogenesis.
Inescapable Need for Neutrophils as Mediators of Cellular Innate Immunity to Acute
Pseudomonas aeruginosa
Pneumonia
Authors: Koh Andrew Y. , Priebe Gregory P. , Ray Christopher , Van Rooijen Nico , Pier Gerald B. 
Published: October 2009

ABSTRACT

Pseudomonas aeruginosa
is a leading cause of pneumonia, and many components of the innate immune system have been proposed to exert important effects in preventing lung infection. However, a vigorous experimental system to identify an overriding, key effector mediating innate immunity to lung infection has not been utilized. As many of the important components of innate immunity are involved in recruitment and activation of polymorphonuclear neutrophils (PMNs) to infected tissues, we hypothesized that the cells and factors needed for their proper recruitment to the lung comprised the major mediators of innate immunity. In neutropenic mice, intranasal (i.n.) doses of
P. aeruginosa
as low as 10 to 100 CFU/mouse produced a fatal lung infection, compared with 10
7
to >10
8
CFU for nonneutropenic mice. There was only a very modest increased mortality in mice lacking mature lymphocytes and no increased mortality in mice depleted of alveolar macrophages when administered i.n.
P. aeruginosa
. Recombinant mouse granulocyte colony-stimulating factor increased survival of neutropenic mice after i.n.
P. aeruginosa
inoculation. MyD88
−/−
mice, which cannot recruit PMNs to the lungs, were highly susceptible to fatal
P. aeruginosa
lung infection, with bacterial doses of <120 CFU being lethal. Activation of a MyD88-independent pathway for PMN recruitment to the lungs in MyD88
−/−
mice resulted in enhanced protection against
P. aeruginosa
lung infection. Overall, in the absence of PMNs, mice cannot resist
P. aeruginosa
lung infection from extremely small bacterial doses. There is an inescapable requirement for local PMN recruitment and activation to mediate innate immunity to
P. aeruginosa
lung infection.
Leptospira interrogans
Binds to Human Cell Surface Receptors Including Proteoglycans
Authors: Breiner Deborah D. , Fahey Mark , Salvador Ryan , Novakova Jana , Coburn Jenifer 
Published: October 2009

ABSTRACT

Leptospirosis is a global public health problem, primarily in the tropical developing world. The pathogenic mechanisms of the causative agents, several members of the genus
Leptospira
, have been underinvestigated. The exception to this trend has been the demonstration of the binding of pathogenic leptospires to the extracellular matrix (ECM) and its components. In this work, interactions of
Leptospira interrogans
bacteria with mammalian cells, rather than the ECM, were examined. The bacteria bound more efficiently to the cells than to the ECM, and a portion of this cell-binding activity was attributable to attachment to glycosaminoglycan (GAG) chains of proteoglycans (PGs). Chondroitin sulfate B PGs appeared to be the primary targets of
L. interrogans
attachment, while heparan sulfate PGs were much less important. Inhibition of GAG/PG-mediated attachment resulted in partial inhibition of bacterial attachment, suggesting that additional receptors for
L. interrogans
await identification. GAG binding may participate in the pathogenesis of leptospirosis within the host animal. In addition, because GAGs are expressed on the luminal aspects of epithelial cells in the proximal tubules of the kidneys, this activity may play a role in targeting the bacteria to this critical site. Because GAGs are shed in the urine, GAG binding may also be important for transmission to new hosts through the environment.
Enterococcus faecalis
Capsular Polysaccharide Serotypes C and D and Their Contributions to Host Innate Immune Evasion
Authors: Thurlow Lance R. , Thomas Vinai Chittezham , Fleming Sherry D. , Hancock Lynn E. 
Published: October 2009

ABSTRACT

It has become increasingly difficult to treat infections caused by
Enterococcus faecalis
due to its high levels of intrinsic and acquired antibiotic resistance. However, few studies have explored the mechanisms that
E. faecalis
employs to circumvent the host innate immune response and establish infection. Capsular polysaccharides are important virulence factors that are associated with innate immune evasion. We demonstrate, using cultured macrophages (RAW 264.7), that capsule-producing
E. faecalis
strains of either serotype C or D are more resistant to complement-mediated opsonophagocytosis than unencapsulated strains. We show that differences in opsonophagocytosis are not due to variations in C3 deposition but are due to the ability of capsule to mask bound C3 from detection on the surface of
E. faecalis
. Similarly,
E. faecalis
capsule masks lipoteichoic acid from detection, which correlates with decreased tumor necrosis factor alpha production by cultured macrophages in the presence of encapsulated strains compared to that in the presence of unencapsulated strains. Our studies confirm the important role of the capsule as a virulence factor of
E. faecalis
and provide several mechanisms by which the presence of the capsule influences evasion of the innate immune response and suggest that the capsule could be a potential target for developing alternative therapies to treat
E. faecalis
infections.
Instantaneous Within-Patient Diversity of
Pseudomonas aeruginosa
Quorum-Sensing Populations from Cystic Fibrosis Lung Infections
Authors: Wilder Cara N. , Allada Gopal , Schuster Martin 
Published: October 2009

ABSTRACT

In the opportunistic pathogen
Pseudomonas aeruginosa
, acyl-homoserine lactone (acyl-HSL) quorum sensing (QS) regulates biofilm formation and expression of many extracellular virulence factors. Curiously, QS-deficient variants, often carrying mutations in the central QS regulator LasR, are frequently isolated from infections, particularly from cystic fibrosis (CF) lung infections. Very little is known about the proportion and diversity of these QS variants in individual infections. Such information is desirable to better understand the selective forces that drive the evolution of QS phenotypes, including social cheating and innate (nonsocial) benefits. To obtain insight into the instantaneous within-patient diversity of QS, we assayed a panel of 135 concurrent
P. aeruginosa
isolates from eight different adult CF patients (9 to 20 isolates per patient) for various QS-controlled phenotypes. Most patients contained complex mixtures of QS-proficient and -deficient isolates. Among all patients, deficiency in individual phenotypes ranged from 0 to about 90%. Acyl-HSL, sequencing, and complementation analyses of variants with global loss-of-function phenotypes revealed dependency upon the central QS circuitry genes
lasR
,
lasI
, and
rhlI
. Deficient and proficient isolates were clonally related, implying evolution from a common ancestor in vivo. Our results show that the diversity of QS types is high within and among patients, suggesting diverse selection pressures in the CF lung. A single selective mechanism, be it of a social or nonsocial nature, is unlikely to account for such heterogeneity. The observed diversity also shows that conclusions about the properties of
P. aeruginosa
QS populations in individual CF infections cannot be drawn from the characterization of one or a few selected isolates.
Development and Application of New Mouse Models To Study the Pathogenesis of
Clostridium perfringens
Type C Enterotoxemias
Authors: Uzal Francisco A. , Saputo Juliann , Sayeed Sameera , Vidal Jorge E. , Fisher Derek J. , Poon Rachael , Adams Vicki , Fernandez-Miyakawa Mariano E. , Rood Julian I. , McClane Bruce A. 
Published: October 2009

ABSTRACT

Clostridium perfringens
type C isolates cause enterotoxemias and enteritis in humans and livestock. While the major disease signs and lesions of type C disease are usually attributed to beta toxin (CPB), these bacteria typically produce several different lethal toxins. Since understanding of disease pathogenesis and development of improved vaccines is hindered by the lack of small animal models mimicking the lethality caused by type C isolates, in this study we developed two mouse models of
C. perfringens
type C-induced lethality. When inoculated into BALB/c mice by intragastric gavage, 7 of 14 type C isolates were lethal, whereas when inoculated intraduodenally, these strains were all lethal in these mice. Clinical signs in intragastrically and intraduodenally challenged mice were similar and included respiratory distress, abdominal distension, and neurological alterations. At necropsy, the small, and occasionally the large, intestine was dilated and gas filled in most mice developing a clinical response. Histological changes in the gut were relatively mild, consisting of attenuation of the mucosa with villus blunting. Inactivation of the CPB-encoding gene rendered the highly virulent type C strain CN3685 avirulent in the intragastric model and nearly nonlethal in the intraduodenal model. In contrast, inactivation of the genes encoding alpha toxin and perfringolysin O only slightly decreased the lethality of CN3685. Mice could be protected against lethality by intravenous passive immunization with a CPB antibody prior to intragastric challenge. This study proves that CPB is a major contributor to the systemic effects of type C infections and provides new mouse models for investigating the pathogenesis of type C-induced lethality.