scholarly journals In VivoGene Essentiality and Metabolism inBordetella pertussis

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Laura A. Gonyar ◽  
Patrick E. Gelbach ◽  
Dennis G. McDuffie ◽  
Alexander F. Koeppel ◽  
Qing Chen ◽  
...  
Keyword(s):  
Respiratory Tract ◽  
Murine Model ◽  
Whooping Cough ◽  
Critical Role ◽  
Respiratory Illness ◽  
Human Respiratory Tract ◽  
Content Type ◽  
Gene Essentiality

ABSTRACTBordetella pertussisis the causative agent of whooping cough, a serious respiratory illness affecting children and adults, associated with prolonged cough and potential mortality. Whooping cough has reemerged in recent years, emphasizing a need for increased knowledge of basic mechanisms ofB. pertussisgrowth and pathogenicity. While previous studies have provided insight intoin vitrogene essentiality of this organism, very little is known aboutin vivogene essentiality, a critical gap in knowledge, sinceB. pertussishas no previously identified environmental reservoir and is isolated from human respiratory tract samples. We hypothesize that the metabolic capabilities ofB. pertussisare especially tailored to the respiratory tract and that many of the genes involved inB. pertussismetabolism would be required to establish infectionin vivo. In this study, we generated a diverse library of transposon mutants and then used it to probe gene essentialityin vivoin a murine model of infection. Using the CON-ARTIST pipeline, 117 genes were identified as conditionally essential at 1 day postinfection, and 169 genes were identified as conditionally essential at 3 days postinfection. Most of the identified genes were associated with metabolism, and we utilized two existing genome-scale metabolic network reconstructions to probe the effects of individual essential genes on biomass synthesis. This analysis suggested a critical role for glucose metabolism and lipooligosaccharide biosynthesisin vivo. This is the first genome-wide evaluation ofin vivogene essentiality inB. pertussisand provides tools for future exploration.IMPORTANCEOur study describes the firstin vivotransposon sequencing (Tn-seq) analysis ofB. pertussisand identifies genes predicted to be essential forin vivogrowth in a murine model of intranasal infection, generating key resources for future investigations intoB. pertussispathogenesis and vaccine design.

scholarly journals Quantification of the Adenylate Cyclase Toxin of Bordetella pertussisIn Vitroand during Respiratory Infection

2013 ◽  
Vol 81 (5) ◽  
pp. 1390-1398 ◽  
Author(s):  
Joshua C. Eby ◽  
Mary C. Gray ◽  
Jason M. Warfel ◽  
Christopher D. Paddock ◽  
Tara F. Jones ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Respiratory Tract ◽  
Whooping Cough ◽  
Extensive Study ◽  
Target Cells ◽  
Human Infants ◽  
Content Type ◽  
Adenylate Cyclase Toxin

ABSTRACTWhooping cough results from infection of the respiratory tract withBordetella pertussis, and the secreted adenylate cyclase toxin (ACT) is essential for the bacterium to establish infection. Despite extensive study of the mechanism of ACT cytotoxicity and its effects over a range of concentrationsin vitro, ACT has not been observed or quantifiedin vivo, and thus the concentration of ACT at the site of infection is unknown. The recently developed baboon model of infection mimics the prolonged cough and transmissibility of pertussis, and we hypothesized that measurement of ACT in nasopharyngeal washes (NPW) from baboons, combined with human andin vitrodata, would provide an estimate of the ACT concentration in the airway during infection. NPW contained up to ∼108CFU/mlB. pertussisand 1 to 5 ng/ml ACT at the peak of infection. Nasal aspirate specimens from two human infants with pertussis contained bacterial concentrations similar to those in the baboons, with 12 to 20 ng/ml ACT. When ∼108CFU/ml of a laboratory strain ofB. pertussiswas culturedin vitro, ACT production was detected in 60 min and reached a plateau of ∼60 ng/ml in 6 h. Furthermore, when bacteria were brought into close proximity to target cells by centrifugation, intoxication was increased 4-fold. Collectively, these data suggest that at the bacterium-target cell interface during infection of the respiratory tract, the concentration of ACT can exceed 100 ng/ml, providing a reference point for future studies of ACT and pertussis pathogenesis.

scholarly journals Dissecting the Structure-Function Relationship of a Fungicidal Peptide Derived from the Constant Region of Human Immunoglobulins

2016 ◽  
Vol 60 (4) ◽  
pp. 2435-2442 ◽  
Author(s):  
Tecla Ciociola ◽  
Thelma A. Pertinhez ◽  
Laura Giovati ◽  
Martina Sperindè ◽  
Walter Magliani ◽  
...  
Keyword(s):  
Self Assembly ◽  
Galleria Mellonella ◽  
Critical Role ◽  
Yeast Cells ◽  
Therapeutic Effects ◽  
Constant Region ◽  
Content Type ◽  
Complementarity Determining Regions

ABSTRACTSynthetic peptides encompassing sequences related to the complementarity-determining regions of antibodies or derived from their constant region (Fc peptides) were proven to exert differential antimicrobial, antiviral, antitumor, and/or immunomodulatory activitiesin vitroand/orin vivo, regardless of the specificity and isotype of the parental antibody. Alanine substitution derivatives of these peptides exhibited unaltered, increased, or decreased candidacidal activitiesin vitro. The bioactive IgG-derived Fc N10K peptide (NQVSLTCLVK) spontaneously self-assembles, a feature previously recognized as relevant for the therapeutic activity of another antibody-derived peptide. We evaluated the contribution of each residue to the peptide self-assembling capability by circular-dichroism spectroscopy. The interaction of the N10K peptide and its derivatives withCandida albicanscells was studied by confocal, transmission, and scanning electron microscopy. The apoptosis and autophagy induction profiles in yeast cells treated with the peptides were evaluated by flow cytometry, and the therapeutic efficacy against candidal infection was studied in aGalleria mellonellamodel. Overall, the results indicate a critical role for some residues in the self-assembly process and a correlation of that capability with the candidacidal activities of the peptidesin vitroand their therapeutic effectsin vivo.

scholarly journals Combined Stimulation of Toll-Like Receptor 5 and NOD1 Strongly Potentiates Activity of NF-κB, Resulting in Enhanced Innate Immune Reactions and Resistance to Salmonella enterica Serovar Typhimurium Infection

2013 ◽  
Vol 81 (10) ◽  
pp. 3855-3864 ◽  
Author(s):  
Amir I. Tukhvatulin ◽  
Ilya I. Gitlin ◽  
Dmitry V. Shcheblyakov ◽  
Natalia M. Artemicheva ◽  
Lyudmila G. Burdelya ◽  
...  
Keyword(s):  
Critical Role ◽  
Immune Defense ◽  
Innate Immune ◽  
Microbial Infection ◽  
Immune Reactions ◽  
Content Type ◽  
Essential Components ◽  
Stimulation Of

ABSTRACTPathogen recognition receptors (PRRs) are essential components of host innate immune systems that detect specific conserved pathogen-associated molecular patterns (PAMPs) presented by microorganisms. Members of two families of PRRs, transmembrane Toll-like receptors (TLRs 1, 2, 4, 5, and 6) and cytosolic NOD receptors (NOD1 and NOD2), are stimulated upon recognition of various bacterial PAMPs. Such stimulation leads to induction of a number of immune defense reactions, mainly triggered via activation of the transcription factor NF-κB. While coordination of responses initiated via different PRRs sensing multiple PAMPS present during an infection makes clear biological sense for the host, such interactions have not been fully characterized. Here, we demonstrate that combined stimulation of NOD1 and TLR5 (as well as other NOD and TLR family members) strongly potentiates activity of NF-κB and induces enhanced levels of innate immune reactions (e.g., cytokine production) bothin vitroandin vivo. Moreover, we show that an increased level of NF-κB activity plays a critical role in formation of downstream responses. In live mice, synergy between these receptors resulting in potentiation of NF-κB activity was organ specific, being most prominent in the gastrointestinal tract. Coordinated activity of NOD1 and TLR5 significantly increased protection of mice against enteroinvasiveSalmonellainfection. Obtained results suggest that cooperation of NOD and TLR receptors is important for effective responses to microbial infectionin vivo.

scholarly journals Prediction of Aerosol Deposition in the Human Respiratory Tract via Computational Models: A Review with Recent Updates

Atmosphere ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 137 ◽  
Author(s):  
Vu Khac Hoang Bui ◽  
Ju-Young Moon ◽  
Minhe Chae ◽  
Duckshin Park ◽  
Young-Chul Lee
Keyword(s):  
Respiratory Tract ◽  
Particle Deposition ◽  
Computational Models ◽  
Aerosol Particles ◽  
Three Dimensional ◽  
Aerosol Deposition ◽  
Research Area ◽  
Human Respiratory Tract

The measurement of deposited aerosol particles in the respiratory tract via in vivo and in vitro approaches is difficult due to those approaches’ many limitations. In order to overcome these obstacles, different computational models have been developed to predict the deposition of aerosol particles inside the lung. Recently, some remarkable models have been developed based on conventional semi-empirical models, one-dimensional whole-lung models, three-dimensional computational fluid dynamics models, and artificial neural networks for the prediction of aerosol-particle deposition with a high accuracy relative to experimental data. However, these models still have some disadvantages that should be overcome shortly. In this paper, we take a closer look at the current research trends as well as the future directions of this research area.

scholarly journals Efficacy of Humanized High-Dose Meropenem, Cefepime, and Levofloxacin against Enterobacteriaceae Isolates Producing Verona Integron-Encoded Metallo-β-Lactamase (VIM) in a Murine Thigh Infection Model

2015 ◽  
Vol 59 (11) ◽  
pp. 7145-7147 ◽  
Author(s):  
Islam M. Ghazi ◽  
Jared L. Crandon ◽  
Emil P. Lesho ◽  
Patrick McGann ◽  
David P. Nicolau
Keyword(s):  
Murine Model ◽  
Infection Model ◽  
High Dose ◽  
Content Type ◽  
High Mics

ABSTRACTWe aimed to describe thein vivoactivity of humanized pharmacokinetic exposures of meropenem and comparators against Verona integron-encoded metallo-β-lactamase (MBL) (VIM)-producingEnterobacteriaceaein a murine model. Levofloxacin activity was predicted by its MIC, and cefepime activity displayed variability, whereas meropenem produced a >1 log CFU reduction against all isolates despite high MICs indicative of resistance. Our results suggest that despitein vitroresistance, high-dose meropenem may be a possible option against infections caused byEnterobacteriaceaeproducing MBL-type carbapenemases.

scholarly journals Enantiomers of Nifurtimox Do Not Exhibit Stereoselective Anti-Trypanosoma cruzi Activity, Toxicity, or Pharmacokinetic Properties

2015 ◽  
Vol 59 (6) ◽  
pp. 3645-3647 ◽  
Author(s):  
Carolina B. Moraes ◽  
Karen L. White ◽  
Stéphanie Braillard ◽  
Catherine Perez ◽  
Junghyun Goo ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Murine Model ◽  
Therapeutic Benefit ◽  
Racemic Mixture ◽  
Content Type ◽  
Pharmacokinetic Properties

ABSTRACTWith the aim of improving the available drugs for the treatment of Chagas disease, individual enantiomers of nifurtimox were characterized. The results indicate that the enantiomers are equivalent in theirin vitroactivity against a panel ofTrypanosoma cruzistrains;in vivoefficacy in a murine model of Chagas disease;in vitrotoxicity and absorption, distribution, metabolism, and excretion characteristics; andin vivopharmacokinetic properties. There is unlikely to be any therapeutic benefit of an individual nifurtimox enantiomer over the racemic mixture.

scholarly journals Macrolones Are a Novel Class of Macrolide Antibiotics Active against Key Resistant Respiratory PathogensIn VitroandIn Vivo

2016 ◽  
Vol 60 (9) ◽  
pp. 5337-5348 ◽  
Author(s):  
Hana Čipčić Paljetak ◽  
Donatella Verbanac ◽  
Jasna Padovan ◽  
Miroslava Dominis-Kramarić ◽  
Željko Kelnerić ◽  
...  
Keyword(s):  
Murine Model ◽  
Bacterial Resistance ◽  
Volume Of Distribution ◽  
Macrolide Antibiotics ◽  
Respiratory Pathogens ◽  
Protein Synthesis Inhibitors ◽  
Content Type ◽  
Tract Infections

ABSTRACTAs we face an alarming increase in bacterial resistance to current antibacterial chemotherapeutics, expanding the available therapeutic arsenal in the fight against resistant bacterial pathogens causing respiratory tract infections is of high importance. The antibacterial potency of macrolones, a novel class of macrolide antibiotics, against key respiratory pathogens was evaluatedin vitroandin vivo. MIC values againstStreptococcus pneumoniae,Streptococcus pyogenes,Staphylococcus aureus, andHaemophilus influenzaestrains sensitive to macrolide antibiotics and with defined macrolide resistance mechanisms were determined. The propensity of macrolones to induce the expression of inducibleermgenes was tested by the triple-disk method and incubation in the presence of subinhibitory concentrations of compounds.In vivoefficacy was assessed in a murine model ofS. pneumoniae-induced pneumonia, and pharmacokinetic (PK) profiles in mice were determined. Thein vitroantibacterial profiles of macrolones were superior to those of marketed macrolide antibiotics, including the ketolide telithromycin, and the compounds did not induce the expression of inducibleermgenes. They acted as typical protein synthesis inhibitors in anEscherichia colitranscription/translation assay. Macrolones were characterized by low to moderate systemic clearance, a large volume of distribution, a long half-life, and low oral bioavailability. They were highly efficacious in a murine model of pneumonia after intraperitoneal application even against anS. pneumoniaestrain with constitutive resistance to macrolide-lincosamide-streptogramin B antibiotics. Macrolones are the class of macrolide antibiotics with an outstanding antibacterial profile and reasonable PK parameters resulting in goodin vivoefficacy.

scholarly journals Streptococcus pyogenes Arginine and Citrulline Catabolism Promotes Infection and Modulates Innate Immunity

2013 ◽  
Vol 82 (1) ◽  
pp. 233-242 ◽  
Author(s):  
Zachary T. Cusumano ◽  
Michael E. Watson ◽  
Michael G. Caparon
Keyword(s):  
Nitric Oxide ◽  
Innate Immunity ◽  
Streptococcus Pyogenes ◽  
Murine Model ◽  
Acid Stress ◽  
Arginine Deiminase ◽  
Content Type ◽  
Cutaneous Tissue

ABSTRACTA bacterium's ability to acquire nutrients from its host during infection is an essential component of pathogenesis. For the Gram-positive pathogenStreptococcus pyogenes, catabolism of the amino acid arginine via the arginine deiminase (ADI) pathway supplements energy production and provides protection against acid stressin vitro. Its expression is enhanced in murine models of infection, suggesting an important rolein vivo. To gain insight into the function of the ADI pathway in pathogenesis, the virulence of mutants defective in each of its enzymes was examined. Mutants unable to use arginine (ΔArcA) or citrulline (ΔArcB) were attenuated for carriage in a murine model of asymptomatic mucosal colonization. However, in a murine model of inflammatory infection of cutaneous tissue, the ΔArcA mutant was attenuated but the ΔArcB mutant was hyperattenuated, revealing an unexpected tissue-specific role for citrulline metabolism in pathogenesis. When mice defective for the arginine-dependent production of nitric oxide (iNOS−/−) were infected with the ΔArcA mutant, cutaneous virulence was rescued, demonstrating that the ability ofS. pyogenesto utilize arginine was dispensable in the absence of nitric oxide-mediated innate immunity. This work demonstrates the importance of arginine and citrulline catabolism and suggests a novel mechanism of virulence by whichS. pyogenesuses its metabolism to modulate innate immunity through depletion of an essential host nutrient.

scholarly journals Histone Chaperone Nap1 Is a Major Regulator of Histone H2A-H2B Dynamics at the InducibleGALLocus

2016 ◽  
Vol 36 (8) ◽  
pp. 1287-1296 ◽  
Author(s):  
Xu Chen ◽  
Sheena D'Arcy ◽  
Catherine A. Radebaugh ◽  
Daniel D. Krzizike ◽  
Holli A. Giebler ◽  
...  
Keyword(s):  
Critical Role ◽  
Histone Chaperones ◽  
Histone H2a ◽  
Nucleosome Assembly ◽  
Content Type ◽  
Nucleosome Assembly Protein ◽  
Nucleosome Assembly Protein 1 ◽  
Histone Exchange

Histone chaperones, like nucleosome assembly protein 1 (Nap1), play a critical role in the maintenance of chromatin architecture. Here, we use theGALlocus inSaccharomyces cerevisiaeto investigate the influence of Nap1 on chromatin structure and histone dynamics during distinct transcriptional states. When theGALlocus is not expressed, cells lacking Nap1 show an accumulation of histone H2A-H2B but not histone H3-H4 at this locus. Excess H2A-H2B interacts with the linker DNA between nucleosomes, and the interaction is independent of the inherent DNA-binding affinity of H2A-H2B for these particular sequences as measuredin vitro. When theGALlocus is transcribed, excess H2A-H2B is reversed, and levels of all chromatin-bound histones are depleted in cells lacking Nap1. We developed anin vivosystem to measure histone exchange at theGALlocus and observed considerable variability in the rate of exchange across the locus in wild-type cells. We recapitulate this variability within vitronucleosome reconstitutions, which suggests a contribution of DNA sequence to histone dynamics. We also find that Nap1 is required for transcription-dependent H2A-H2B exchange. Altogether, these results indicate that Nap1 is essential for maintaining proper chromatin composition and modulating the exchange of H2A-H2Bin vivo.

scholarly journals Toll-Like Receptor 9 Signaling in Dendritic Cells Regulates Neutrophil Recruitment to Inflammatory Foci following Leishmania infantum Infection

2015 ◽  
Vol 83 (12) ◽  
pp. 4604-4616 ◽  
Author(s):  
Laís Sacramento ◽  
Silvia C. Trevelin ◽  
Manuela S. Nascimento ◽  
Djalma S. Lima-Jùnior ◽  
Diego L. Costa ◽  
...  
Keyword(s):  
Leishmania Infantum ◽  
Critical Role ◽  
Costimulatory Molecule ◽  
Protozoan Parasite ◽  
Neutrophil Recruitment ◽  
Content Type ◽  
Target Organs ◽  
Myd88 Signaling

Leishmania infantumis a protozoan parasite that causes visceral leishmaniasis (VL). This infection triggers dendritic cell (DC) activation through the recognition of microbial products by Toll-like receptors (TLRs). Among the TLRs, TLR9 is required for DC activation by differentLeishmaniaspecies. We demonstrated that TLR9 is upregulatedin vitroandin vivoduring infection. We show that C57BL/6 mice deficient in TLR9 expression (TLR9−/−mice) are more susceptible to infection and display higher parasite numbers in the spleen and liver. The increased susceptibility of TLR9−/−mice was due to the impaired recruitment of neutrophils to the infection foci associated with reduced levels of neutrophil chemoattractants released by DCs in the target organs. Moreover, both Th1 and Th17 cells were also committed in TLR9−/−mice. TLR9-dependent neutrophil recruitment is mediated via the MyD88 signaling pathway but is TIR domain-containing adapter-inducing interferon beta (TRIF) independent. Furthermore,L. infantumfailed to activate both plasmacytoid and myeloid DCs from TLR9−/−mice, which presented reduced surface costimulatory molecule expression and chemokine release. Interestingly, neutrophil chemotaxis was affected bothin vitroandin vivowhen DCs were derived from TLR9−/−mice. Our results suggest that TLR9 plays a critical role in neutrophil recruitment during the protective response againstL. infantuminfection that could be associated with DC activation.

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