scholarly journals The Yfe and Feo Transporters Are Involved in Microaerobic Growth and Virulence of Yersinia pestis in Bubonic Plague

2012 ◽  
Vol 80 (11) ◽  
pp. 3880-3891 ◽  
Author(s):  
Jacqueline D. Fetherston ◽  
Ildefonso Mier ◽  
Helena Truszczynska ◽  
Robert D. Perry
Keyword(s):  
Yersinia Pestis ◽  
Double Mutant ◽  
Lethal Dose ◽  
Growth Conditions ◽  
Transport Systems ◽  
Growth Defect ◽  
Dependent Manner ◽  
Bubonic Plague ◽  
Pneumonic Plague ◽  
Content Type

ABSTRACTThe Yfe/Sit and Feo transport systems are important for the growth of a variety of bacteria. InYersinia pestis, single mutations in eitheryfeorfeoresult in reduced growth under static (limited aeration), iron-chelated conditions, while ayfe feodouble mutant has a more severe growth defect. These growth defects were not observed when bacteria were grown under aerobic conditions or in strains capable of producing the siderophore yersiniabactin (Ybt) and the putative ferrous transporter FetMP. BothfetPand a downstream locus (flpforfetlinkedphenotype) were required for growth of ayfe feo ybtmutant under static, iron-limiting conditions. AnfeoBmutation alone had no effect on the virulence ofY. pestisin either bubonic or pneumonic plague models. Anfeo yfedouble mutant was still fully virulent in a pneumonic plague model but had an ∼90-fold increase in the 50% lethal dose (LD50) relative to the Yfe+Feo+parent strain in a bubonic plague model. Thus, Yfe and Feo, in addition to Ybt, play an important role in the progression of bubonic plague. Finally, we examined the factors affecting the expression of thefeooperon inY. pestis. Under static growth conditions, theY. pestis feo::lacZfusion was repressed by iron in a Fur-dependent manner but not in cells grown aerobically. Mutations infeoC,fnr,arcA,oxyR, orrstABhad no significant effect on transcription of theY. pestis feopromoter. Thus, the factor(s) that prevents repression by Fur under aerobic growth conditions remains to be identified.

scholarly journals Combinational Deletion of Three Membrane Protein-Encoding Genes Highly Attenuates Yersinia pestis while Retaining Immunogenicity in a Mouse Model of Pneumonic Plague

2015 ◽  
Vol 83 (4) ◽  
pp. 1318-1338 ◽  
Author(s):  
Bethany L. Tiner ◽  
Jian Sha ◽  
Michelle L. Kirtley ◽  
Tatiana E. Erova ◽  
Vsevolod L. Popov ◽  
...  
Keyword(s):  
Mouse Model ◽  
Yersinia Pestis ◽  
Double Mutant ◽  
Lethal Dose ◽  
Bactericidal Effect ◽  
Alveolar Epithelial Cells ◽  
Triple Mutant ◽  
Pneumonic Plague ◽  
Content Type

Previously, we showed that deletion of genes encoding Braun lipoprotein (Lpp) and MsbB attenuatedYersinia pestisCO92 in mouse and rat models of bubonic and pneumonic plague. While Lpp activates Toll-like receptor 2, the MsbB acyltransferase modifies lipopolysaccharide. Here, we deleted theailgene (encoding theattachment-invasionlocus) from wild-type (WT) strain CO92 or itslppsingle and ΔlppΔmsbBdouble mutants. While the Δailsingle mutant was minimally attenuated compared to the WT bacterium in a mouse model of pneumonic plague, the ΔlppΔaildouble mutant and the ΔlppΔmsbBΔailtriple mutant were increasingly attenuated, with the latter being unable to kill mice at a 50% lethal dose (LD50) equivalent to 6,800 LD50s of WT CO92. The mutant-infected animals developed balanced TH1- and TH2-based immune responses based on antibody isotyping. The triple mutant was cleared from mouse organs rapidly, with concurrent decreases in the production of various cytokines and histopathological lesions. When surviving animals infected with increasing doses of the triple mutant were subsequently challenged on day 24 with the bioluminescent WT CO92 strain (20 to 28 LD50s), 40 to 70% of the mice survived, with efficient clearing of the invading pathogen, as visualized in real time byin vivoimaging. The rapid clearance of the triple mutant, compared to that of WT CO92, from animals was related to the decreased adherence and invasion of human-derived HeLa and A549 alveolar epithelial cells and to its inability to survive intracellularly in these cells as well as in MH-S murine alveolar and primary human macrophages. An early burst of cytokine production in macrophages elicited by the triple mutant compared to WT CO92 and the mutant's sensitivity to the bactericidal effect of human serum would further augment bacterial clearance. Together, deletion of theailgene from the ΔlppΔmsbBdouble mutant severely attenuatedY. pestisCO92 to evoke pneumonic plague in a mouse model while retaining the required immunogenicity needed for subsequent protection against infection.

scholarly journals Intranasal Administration of an Inactivated Yersinia pestis Vaccine with Interleukin-12 Generates Protective Immunity against Pneumonic Plague

2011 ◽  
Vol 18 (11) ◽  
pp. 1925-1935 ◽  
Author(s):  
Devender Kumar ◽  
Girish Kirimanjeswara ◽  
Dennis W. Metzger
Keyword(s):  
Yersinia Pestis ◽  
Protective Efficacy ◽  
Bacterial Colonization ◽  
Lethal Dose ◽  
Interleukin 12 ◽  
Cell Vaccine ◽  
Mouse Serum ◽  
Immune Mouse ◽  
Pneumonic Plague ◽  
Content Type

ABSTRACTInhalation ofYersinia pestiscauses pneumonic plague, which rapidly progresses to death. A previously licensed killed whole-cell vaccine is presently unavailable due to its reactogenicity and inconclusive evidence of efficacy. The present study now shows that vaccination intranasally (i.n.) with inactivatedY. pestisCO92 (iYp) adjuvanted with interleukin-12 (IL-12) followed by an i.n. challenge with a lethal dose ofY. pestisCO92 prevented bacterial colonization and protected 100% of mice from pneumonic plague. Survival of the vaccinated mice correlated with levels of systemic and lung antibodies, reduced pulmonary pathology and proinflammatory cytokines, and the presence of lung lymphoid cell aggregates. Protection against pneumonic plague was partially dependent upon Fc receptors and could be transferred to naïve mice with immune mouse serum. On the other hand, protection was not dependent upon complement, and following vaccination, depletion of CD4 and/or CD8 T cells before challenge did not affect survival. In summary, the results demonstrate the safety, immunogenicity, and protective efficacy of i.n. administered iYp plus IL-12 in a mouse model of pneumonic plague.

scholarly journals Characterization of a Cynomolgus Macaque Model of Pneumonic Plague for Evaluation of Vaccine Efficacy

2015 ◽  
Vol 22 (9) ◽  
pp. 1070-1078 ◽  
Author(s):  
Patricia Fellows ◽  
Jessica Price ◽  
Shannon Martin ◽  
Karen Metcalfe ◽  
Robert Krile ◽  
...  
Keyword(s):  
Lethal Dose ◽  
Clinical Signs ◽  
Cynomolgus Macaque ◽  
Vaccine Dose ◽  
Dose Schedule ◽  
Dependent Manner ◽  
Pneumonic Plague ◽  
Content Type ◽  
Antibody Titers ◽  
Time To Onset

ABSTRACTThe efficacy of a recombinant plague vaccine (rF1V) was evaluated in cynomolgus macaques (CMs) to establish the relationship among vaccine doses, antibody titers, and survival following an aerosol challenge with a lethal dose ofYersinia pestisstrain Colorado 92. CMs were vaccinated with a range of rF1V doses on a three-dose schedule (days 0, 56, and 121) to provide a range of survival outcomes. The humoral immune response following vaccination was evaluated with anti-rF1, anti-rV, and anti-rF1V bridge enzyme-linked immunosorbent assays (ELISAs). Animals were challenged via aerosol exposure on day 149. Vaccine doses and antibody responses were each significantly associated with the probability of CM survival (P< 0.0001). Vaccination also decreased signs of pneumonic plague in a dose-dependent manner. There were statistically significant correlations between the vaccine dose and the time to onset of fever (P< 0.0001), the time from onset of fever to death (P< 0.0001), the time to onset of elevated respiratory rate (P= 0.0003), and the time to onset of decreased activity (P= 0.0251) postinfection in animals exhibiting these clinical signs. Delays in the onset of these clinical signs of disease were associated with larger doses of rF1V. Immunization with ≥12 μg of rF1V resulted in 100% CM survival. Since both the vaccine dose and anti-rF1V antibody titers correlate with survival, rF1V bridge ELISA titers can be used as a correlate of protection.

scholarly journals Inactivation of Peroxiredoxin 6 by the Pla Protease of Yersinia pestis

2015 ◽  
Vol 84 (1) ◽  
pp. 365-374 ◽  
Author(s):  
Daniel L. Zimbler ◽  
Justin L. Eddy ◽  
Jay A. Schroeder ◽  
Wyndham W. Lathem
Keyword(s):  
Yersinia Pestis ◽  
Lung Damage ◽  
Rapid Progression ◽  
Dependent Manner ◽  
Wild Type ◽  
Peroxiredoxin 6 ◽  
Pneumonic Plague ◽  
Content Type

Pneumonic plague represents the most severe form of disease caused byYersinia pestisdue to its ease of transmission, rapid progression, and high mortality rate. TheY. pestisouter membrane Pla protease is essential for the development of pneumonic plague; however, the complete repertoire of substrates cleaved by Pla in the lungs is not known. In this study, we describe a proteomic screen to identify host proteins contained within the bronchoalveolar lavage fluid of mice that are cleaved and/or processed byY. pestisin a Pla-dependent manner. We identified peroxiredoxin 6 (Prdx6), a host factor that contributes to pulmonary surfactant metabolism and lung defense against oxidative stress, as a previously unknown substrate of Pla. Pla cleaves Prdx6 at three distinct sites, and these cleavages disrupt both the peroxidase and phospholipase A2activities of Prdx6. In addition, we found that infection with wild-typeY. pestisreduces the abundance of extracellular Prdx6 in the lungs compared to that after infection with ΔplaY. pestis, suggesting that Pla cleaves Prdx6 in the pulmonary compartment. However, following infection with either wild-type or Δpla Y. pestis, Prdx6-deficient mice exhibit no differences in bacterial burden, host immune response, or lung damage from wild-type mice. Thus, while Pla is able to disrupt Prdx6 functionin vitroand reduce Prdx6 levelsin vivo, the cleavage of Prdx6 has little detectable impact on the progression or outcome of pneumonic plague.

scholarly journals Dual Zinc Transporter Systems in Vibrio cholerae Promote Competitive Advantages over Gut Microbiome

2015 ◽  
Vol 83 (10) ◽  
pp. 3902-3908 ◽  
Author(s):  
Ying Sheng ◽  
Fenxia Fan ◽  
Owen Jensen ◽  
Zengtao Zhong ◽  
Biao Kan ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Gene Clusters ◽  
Bioinformatic Analysis ◽  
Zinc Transporter ◽  
Zinc Homeostasis ◽  
Transport Systems ◽  
Growth Defect ◽  
Dependent Manner ◽  
Content Type ◽  
Transporter Systems

Zinc is an essential trace metal required for numerous cellular processes in all forms of life. In order to maintain zinc homeostasis, bacteria have developed several transport systems to regulate its uptake. In this study, we investigated zinc transport systems in the enteric pathogenVibrio cholerae, the causative agent of cholera. Bioinformatic analysis predicts that two gene clusters, VC2081 to VC2083 (annotated as zinc utilization genesznuABC) and VC2551 to VC2555 (annotated aszinc-regulatedgeneszrgABCDE), are regulated by the putative zinc uptake regulator Zur. Using promoter reporter and biochemical assays, we confirmed that Zur repressesznuABCandzrgABCDEpromoters in a Zn2+-dependent manner. Under Zn2+-limiting conditions, we found that mutations in either theznuABCorzrgABCDEgene cluster affect bacterial growth, withznuABCmutants displaying a more severe growth defect, suggesting that both ZnuABC and ZrgABCDE are involved in Zn2+uptake and that ZnuABC plays the predominant role. Furthermore, we reveal that ZnuABC and ZrgABCDE are important forV. choleraecolonization in both infant and adult mouse models, particularly in the presence of other intestinal microbiota. Collectively, our studies indicate that these two zinc transporter systems play vital roles in maintaining zinc homeostasis duringV. choleraegrowth and pathogenesis.

scholarly journals Interplay of Nitric Oxide Synthase (NOS) and SrrAB in Modulation ofStaphylococcus aureusMetabolism and Virulence

2018 ◽  
Vol 87 (2) ◽  
Author(s):  
Kimberly L. James ◽  
Austin B. Mogen ◽  
Jessica N. Brandwein ◽  
Silvia S. Orsini ◽  
Miranda J. Ridder ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Double Mutant ◽  
Nitrate Assimilation ◽  
Arginine Deiminase ◽  
Growth Defect ◽  
Content Type ◽  
Metabolic Versatility ◽  
Oxide Synthase

ABSTRACTStaphylococcus aureusnitric oxide synthase (saNOS) is a major contributor to virulence, stress resistance, and physiology, yet the specific mechanism(s) by which saNOS intersects with other known regulatory circuits is largely unknown. The SrrAB two-component system, which modulates gene expression in response to the reduced state of respiratory menaquinones, is a positive regulator ofnosexpression. Several SrrAB-regulated genes were also previously shown to be induced in an aerobically respiringnosmutant, suggesting a potential interplay between saNOS and SrrAB. Therefore, a combination of genetic, molecular, and physiological approaches was employed to characterize anos srrABmutant, which had significant reductions in the maximum specific growth rate and oxygen consumption when cultured under conditions promoting aerobic respiration. Thenos srrABmutant secreted elevated lactate levels, correlating with the increased transcription of lactate dehydrogenases. Expression of nitrate and nitrite reductase genes was also significantly enhanced in thenos srrABdouble mutant, and its aerobic growth defect could be partially rescued with supplementation with nitrate, nitrite, or ammonia. Furthermore, elevated ornithine and citrulline levels and highly upregulated expression of arginine deiminase genes were observed in the double mutant. These data suggest that a dual deficiency in saNOS and SrrAB limitsS. aureusto fermentative metabolism, with a reliance on nitrate assimilation and the urea cycle to help fuel energy production. Thenos,srrAB, andnos srrABmutants showed comparable defects in endothelial intracellular survival, whereas thesrrABandnos srrABmutants were highly attenuated during murine sepsis, suggesting that SrrAB-mediated metabolic versatility is dominantin vivo.

scholarly journals Induction of Protective Antiplague Immune Responses by Self-Adjuvanting Bionanoparticles Derived from Engineered Yersinia pestis

2020 ◽  
Vol 88 (5) ◽  
Author(s):  
Xiuran Wang ◽  
Amit K. Singh ◽  
Xiangmin Zhang ◽  
Wei Sun
Keyword(s):  
Immune Responses ◽  
Yersinia Pestis ◽  
Rational Design ◽  
Lipid A ◽  
Vaccine Development ◽  
Lethal Dose ◽  
Outer Membrane Vesicles ◽  
Vector System ◽  
Content Type ◽  
Monophosphoryl Lipid A

ABSTRACT A Yersinia pestis mutant synthesizing an adjuvant form of lipid A (monophosphoryl lipid A, MPLA) displayed increased biogenesis of bacterial outer membrane vesicles (OMVs). To enhance the immunogenicity of the OMVs, we constructed an Asd-based balanced-lethal host-vector system that oversynthesized the LcrV antigen of Y. pestis, raised the amounts of LcrV enclosed in OMVs by the type II secretion system, and eliminated harmful factors like plasminogen activator (Pla) and murine toxin from the OMVs. Vaccination with OMVs containing MPLA and increased amounts of LcrV with diminished toxicity afforded complete protection in mice against subcutaneous challenge with 8 × 105 CFU (80,000 50% lethal dose [LD50]) and intranasal challenge with 5 × 103 CFU (50 LD50) of virulent Y. pestis. This protection was significantly superior to that resulting from vaccination with LcrV/alhydrogel or rF1-V/alhydrogel. At week 4 postimmunization, the OMV-immunized mice showed more robust titers of antibodies against LcrV, Y. pestis whole-cell lysate (YPL), and F1 antigen and more balanced IgG1:IgG2a/IgG2b-derived Th1 and Th2 responses than LcrV-immunized mice. Moreover, potent adaptive and innate immune responses were stimulated in the OMV-immunized mice. Our findings demonstrate that self-adjuvanting Y. pestis OMVs provide a novel plague vaccine candidate and that the rational design of OMVs could serve as a robust approach for vaccine development.

scholarly journals Inhibition of Neutrophil Primary Granule Release during Yersinia pestis Pulmonary Infection

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Kara R. Eichelberger ◽  
Grant S. Jones ◽  
William E. Goldman
Keyword(s):  
Yersinia Pestis ◽  
Critical Role ◽  
Immune Defense ◽  
Calcium Flux ◽  
Human Neutrophils ◽  
Pneumonic Plague ◽  
Content Type ◽  
Neutrophil Degranulation ◽  
Primary Granule ◽  
Granule Release

ABSTRACT Inhalation of Yersinia pestis causes primary pneumonic plague, the most severe manifestation of plague that is characterized by a dramatic neutrophil influx to the lungs. Neutrophils are ineffective during primary pneumonic plague, failing to control Y. pestis growth in the airways. However, the mechanisms by which Y. pestis resists neutrophil killing are incompletely understood. Here, we show that Y. pestis inhibits neutrophil degranulation, an important line of host innate immune defense. We observed that neutrophils from the lungs of mice infected intranasally with Y. pestis fail to release primary granules throughout the course of disease. Using a type III secretion system (T3SS) injection reporter strain, we determined that Y. pestis directly inhibits neutrophil granule release by a T3SS-dependent mechanism. Combinatorial mutant analysis revealed that a Y. pestis strain lacking both effectors YopE and YopH did not inhibit primary granule release and is killed by neutrophils both in vivo and in vitro. Similarly, Y. pestis strains injecting only YopE or YopH are able to inhibit the majority of primary granule release from human neutrophils. We determined that YopE and YopH block Rac2 activation and calcium flux, respectively, to inhibit neutrophil primary granule release in isolated human neutrophils. These results demonstrate that Y. pestis coordinates the inhibition of neutrophil primary granule release through the activities of two distinct effectors, and this inhibition promotes Y. pestis survival during primary pneumonic plague. IMPORTANCE Yersinia pestis is the causative agent of plague and is one of the deadliest human pathogens. The pneumonic form of Y. pestis infection has played a critical role in the severity of both historical and modern plague outbreaks, yet the host-pathogen interactions that govern the lethality of Yersinia pestis pulmonary infections are incompletely understood. Here, we report that Yersinia pestis inhibits neutrophil degranulation during infection, rendering neutrophils ineffective and allowing unrestricted growth of Y. pestis in the lungs. This coordinated inhibition of granule release not only demonstrates the pathogenic benefit of “silencing” lung neutrophils but also reveals specific host processes and pathways that could be manipulated to reduce the severity of primary pneumonic plague.

scholarly journals Posttranscriptional Regulation of the Yersinia pestis Cyclic AMP Receptor Protein Crp and Impact on Virulence

mBio ◽  
2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Wyndham W. Lathem ◽  
Jay A. Schroeder ◽  
Lauren E. Bellows ◽  
Jeremy T. Ritzert ◽  
Jovanka T. Koo ◽  
...  
Keyword(s):  
Cyclic Amp ◽  
Yersinia Pestis ◽  
Posttranscriptional Regulation ◽  
Transcriptional Regulator ◽  
Receptor Protein ◽  
Dependent Manner ◽  
Content Type ◽  
Cyclic Amp Receptor Protein ◽  
Protein Levels ◽  
Posttranscriptional Level

ABSTRACTThe cyclic AMP receptor protein (Crp) is a transcriptional regulator that controls the expression of numerous bacterial genes, usually in response to environmental conditions and particularly by sensing the availability of carbon. In the plague pathogenYersinia pestis, Crp regulates the expression of multiple virulence factors, including components of the type III secretion system and the plasminogen activator protease Pla. The regulation of Crp itself, however, is distinctly different from that found in the well-studiedEscherichia colisystem. Here, we show that at physiological temperatures, the synthesis of Crp inY. pestisis positively regulated at the posttranscriptional level. The loss of the small RNA chaperone Hfq results in decreased Crp protein levels but not in steady-state Crp transcript levels, and this regulatory effect occurs within the 5′ untranslated region (UTR) of the Crp mRNA. The posttranscriptional activation of Crp synthesis is required for the expression ofpla, and decouplingcrpfrom Hfq through the use of an exogenously controlled promoter and 5′ UTR increases Pla protein levels as well as partially rescues the growth defect associated with the loss of Hfq. Finally, we show that both Hfq and the posttranscriptional regulation of Crp contribute to the virulence ofY. pestisduring pneumonic plague. The Hfq-dependent, posttranscriptional regulation of Crp may be specific toYersiniaspecies, and thus our data help explain the dramatic growth and virulence defects associated with the loss of Hfq inY. pestis.IMPORTANCEThe Crp protein is a major transcriptional regulator in bacteria, and its synthesis is tightly controlled to avoid inappropriate induction of the Crp regulon. In this report, we provide the first evidence of Crp regulation in an Hfq-dependent manner at the posttranscriptional level. Our discovery that the synthesis of Crp inYersinia pestisis Hfq dependent adds an additional layer of regulation to catabolite repression in this bacterium. Our work provides a mechanism by which the plague pathogen links not just the sensing of glucose or other carbon sources but also other signals that influence Crp abundance via the expression of small RNAs to the induction of the Crp regulon. In turn, this allowsY. pestisto fine-tune Crp levels to optimize virulence gene expression during plague infection and may allow the bacterium to adapt to its unique environmental niches.

scholarly journals Gr1+ Cells Control Growth of YopM-Negative Yersinia pestis during Systemic Plague

2009 ◽  
Vol 77 (9) ◽  
pp. 3791-3806 ◽  
Author(s):  
Zhan Ye ◽  
Edward J. Kerschen ◽  
Donald A. Cohen ◽  
Alan M. Kaplan ◽  
Nico van Rooijen ◽  
...  
Keyword(s):  
Nk Cells ◽  
Yersinia Pestis ◽  
Nk Cell ◽  
Early Stage ◽  
Lethal Dose ◽  
Cell Types ◽  
Growth Defect ◽  
Protein Toxin ◽  
Inflammatory Monocytes ◽  
Control Growth

ABSTRACT YopM, a protein toxin of Yersinia pestis, is necessary for virulence in a mouse model of systemic plague. We previously reported YopM-dependent natural killer (NK) cell depletion from blood and spleen samples of infected mice. However, in this study we found that infection with Y. pestis KIM5 (YopM+) caused depletion of NK cells in the spleen, but not in the liver, and antibody-mediated ablation of NK cells had no effect on bacterial growth. There was no YopM-associated effect on the percentage of dendritic cells (DCs) or polymorphonuclear leukocytes (PMNs) in the early stage of infection; however, there was a YopM-associated effect on PMN integrity and on the influx of monocytes into the spleen. Ablation of Gr1+ cells caused loss of the growth defect of YopM− Y. pestis in both the liver and spleen. In contrast, ablation of macrophages/DCs inhibited growth of both parent and mutant bacteria, accompanied by significantly fewer lesion sites in the liver. These results point toward PMNs and inflammatory monocytes as major cell types that control growth of YopM− Y. pestis. Infection with fully virulent Y. pestis CO92 and a YopM− derivative by intradermal and intranasal routes showed that the absence of YopM significantly increased the 50% lethal dose only in the intradermal model, suggesting a role for YopM in bubonic plague, in which acute inflammation occurs soon after infection.

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