scholarly journals New Insights into Autoinducer-2 Signaling as a Virulence Regulator in a Mouse Model of Pneumonic Plague

mSphere ◽  
2016 ◽  
Vol 1 (6) ◽  
Author(s):  
Eric C. Fitts ◽  
Jourdan A. Andersson ◽  
Michelle L. Kirtley ◽  
Jian Sha ◽  
Tatiana E. Erova ◽  
...  
Keyword(s):  
Mouse Model ◽  
Yersinia Pestis ◽  
Bacterial Virulence ◽  
Resistant Bacteria ◽  
Antibiotic Resistant Bacteria ◽  
Global Crisis ◽  
Antibiotic Resistant ◽  
Pneumonic Plague ◽  
Content Type ◽  
Autoinducer 2

ABSTRACT Yersinia pestis is the bacterial agent that causes the highly fatal disease plague. The organism represents a significant concern because of its potential use as a bioterror agent, beyond the several thousand naturally occurring human infection cases occurring globally each year. While there has been development of effective antibiotics, the narrow therapeutic window and challenges posed by the existence of antibiotic-resistant strains represent serious concerns. We sought to identify novel virulence factors that could potentially be incorporated into an attenuated vaccine platform or be targeted by novel therapeutics. We show here that a highly conserved quorum-sensing system, autoinducer-2, significantly affected the virulence of Y. pestis in a mouse model of pneumonic plague. We also identified steps in autoinducer-2 signaling which had confounded previous studies and demonstrated the potential for intervention in the virulence mechanism(s) of autoinducer-2. Our findings may have an impact on bacterial pathogenesis research in many other organisms and could result in identifying potential broad-spectrum therapeutic targets to combat antibiotic-resistant bacteria, which represent a global crisis of the 21st century. The Enterobacteriaceae family members, including the infamous Yersinia pestis, the causative agent of plague, have a highly conserved interbacterial signaling system that is mediated by the autoinducer-2 (AI-2) quorum-sensing molecule. The AI-2 system is implicated in regulating various bacterial virulence genes in diverse environmental niches. Deletion of the gene encoding the synthetic enzyme for the AI-2 substrate, luxS, leads to either no significant change or, paradoxically, an increase in in vivo bacterial virulence. We showed that deletion of the rbsA and lsrA genes, components of ABC transport systems that interact with AI-2, synergistically disrupted AI-2 signaling patterns and resulted in a more-than-50-fold decrease in Y. pestis strain CO92 virulence in a stringent pneumonic plague mouse model. Deletion of luxS or lsrK (encoding AI-2 kinase) from the ΔrbsA ΔlsrA background strain or complementation of the ΔrbsA ΔlsrA mutant with the corresponding gene(s) reverted the virulence phenotype to that of the wild-type Y. pestis CO92. Furthermore, the administration of synthetic AI-2 in mice infected with the ΔrbsA ΔlsrA ΔluxS mutant strain attenuated this triple mutant to a virulence phenotype similar to that of the ΔrbsA ΔlsrA strain in a pneumonic plague model. Conversely, the administration of AI-2 to mice infected with the ΔrbsA ΔlsrA ΔluxS ΔlsrK mutant did not rescue animals from lethality, indicating the importance of the AI-2–LsrK axis in regulating bacterial virulence. By performing high-throughput RNA sequencing, the potential role of some AI-2-signaling-regulated genes that modulated bacterial virulence was determined. We anticipate that the characterization of AI-2 signaling in Y. pestis will lead to reexamination of AI-2 systems in other pathogens and that AI-2 signaling may represent a broad-spectrum therapeutic target to combat antibiotic-resistant bacteria, which represent a global crisis of the 21st century. IMPORTANCE Yersinia pestis is the bacterial agent that causes the highly fatal disease plague. The organism represents a significant concern because of its potential use as a bioterror agent, beyond the several thousand naturally occurring human infection cases occurring globally each year. While there has been development of effective antibiotics, the narrow therapeutic window and challenges posed by the existence of antibiotic-resistant strains represent serious concerns. We sought to identify novel virulence factors that could potentially be incorporated into an attenuated vaccine platform or be targeted by novel therapeutics. We show here that a highly conserved quorum-sensing system, autoinducer-2, significantly affected the virulence of Y. pestis in a mouse model of pneumonic plague. We also identified steps in autoinducer-2 signaling which had confounded previous studies and demonstrated the potential for intervention in the virulence mechanism(s) of autoinducer-2. Our findings may have an impact on bacterial pathogenesis research in many other organisms and could result in identifying potential broad-spectrum therapeutic targets to combat antibiotic-resistant bacteria, which represent a global crisis of the 21st century.

scholarly journals Phage Therapy Is Effective in a Mouse Model of Bacterial Equine Keratitis

2016 ◽  
Vol 82 (17) ◽  
pp. 5332-5339 ◽  
Author(s):  
Takaaki Furusawa ◽  
Hidetomo Iwano ◽  
Yutaro Hiyashimizu ◽  
Kazuki Matsubara ◽  
Hidetoshi Higuchi ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Mouse Model ◽  
Phage Therapy ◽  
Host Cells ◽  
Resistant Bacteria ◽  
Broad Host Range ◽  
Bacterial Keratitis ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Content Type

ABSTRACTBacterial keratitis of the horse is mainly caused by staphylococci, streptococci, and pseudomonads. Of these bacteria,Pseudomonas aeruginosasometimes causes rapid corneal corruption and, in some cases, blindness. Antimicrobial resistance can make treatment very difficult. Therefore, new strategies to control bacterial infection are required. A bacteriophage (phage) is a virus that specifically infects and kills bacteria. Since phage often can lyse antibiotic-resistant bacteria because the killing mechanism is different, we examined the use of phage to treat horse bacterial keratitis. We isolatedMyoviridaeorPodoviridaephages, which together have a broad host range. They adsorb efficiently to host bacteria; more than 80% of the ΦR18 phage were adsorbed to host cells after 30 s. In our keratitis mouse model, the administration of phage within 3 h also could kill bacteria and suppress keratitis. A phage multiplicity of infection of 100 times the host bacterial number could kill host bacteria effectively. A cocktail of two phages suppressed bacteria in the keratitis model mouse. These data demonstrated that the phages in this study could completely prevent the keratitis caused byP. aeruginosain a keratitis mouse model. Furthermore, these results suggest that phage may be a more effective prophylaxis for horse keratitis than the current preventive use of antibiotics. Such treatment may reduce the use of antibiotics and therefore antibiotic resistance. Further studies are required to assess phage therapy as a candidate for treatment of horse keratitis.IMPORTANCEAntibiotic-resistant bacteria are emerging all over the world. Bacteriophages have great potential for resolution of this problem. A bacteriophage, or phage, is a virus that infects bacteria specifically. As a novel therapeutic strategy against racehorse keratitis caused byPseudomonas aeruginosa, we propose the application of phages for treatment. Phages isolated in this work hadin vitroeffectiveness for a broad range ofP. aeruginosastrains. Indeed, a great reduction of bacterial proliferation was shown in phage therapy for mouse models ofP. aeruginosakeratitis. Therefore, to reduce antibiotic usage, phage therapy should be investigated and developed further.

scholarly journals Widely Used Benzalkonium Chloride Disinfectants Can Promote Antibiotic Resistance

2018 ◽  
Vol 84 (17) ◽  
Author(s):  
Minjae Kim ◽  
Michael R. Weigand ◽  
Seungdae Oh ◽  
Janet K. Hatt ◽  
Raj Krishnan ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Microbial Communities ◽  
Gene Cloning ◽  
Benzalkonium Chloride ◽  
Efflux Pump ◽  
Resistant Bacteria ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Content Type ◽  
Physiological Adaptations

ABSTRACTWhile the misuse of antibiotics has clearly contributed to the emergence and proliferation of resistant bacterial pathogens, with major health consequences, it remains less clear if the widespread use of disinfectants, such as benzalkonium chlorides (BAC), a different class of biocides than antibiotics, has contributed to this problem. Here, we provide evidence that exposure to BAC coselects for antibiotic-resistant bacteria and describe the underlying genetic mechanisms. After inoculation with river sediment, BAC-fed bioreactors selected for several bacterial taxa, including the opportunistic pathogenPseudomonas aeruginosa, that were more resistant to several antibiotics than their counterparts in a control (no BAC) bioreactor. A metagenomic analysis of the bioreactor microbial communities, confirmed by gene cloning experiments with the derived isolates, suggested that integrative and conjugative elements encoding a BAC efflux pump together with antibiotic resistance genes were responsible for these results. Furthermore, the exposure of theP. aeruginosaisolates to increasing concentrations of BAC selected for mutations inpmrB(polymyxin resistance) and physiological adaptations that contributed to a higher tolerance to polymyxin B and other antibiotics. The physiological adaptations included the overexpression ofmexCD-oprJmultidrug efflux pump genes when BAC was added in the growth medium at subinhibitory concentrations. Collectively, our results demonstrated that disinfectants promote antibiotic resistance via several mechanisms and highlight the need to remediate (degrade) disinfectants in nontarget environments to further restrain the spread of antibiotic-resistant bacteria.IMPORTANCEBenzalkonium chlorides (BAC) are biocides broadly used in disinfectant solutions. Disinfectants are widely used in food processing lines, domestic households, and pharmaceutical products and are typically designed to have a different mode of action than antibiotics to avoid interfering with the use of the latter. Whether exposure to BAC makes bacteria more resistant to antibiotics remains an unresolved issue of obvious practical consequences for public health. Using an integrated approach that combines metagenomics of natural microbial communities with gene cloning experiments with isolates and experimental evolution assays, we show that the widely used benzalkonium chloride disinfectants promote clinically relevant antibiotic resistance. Therefore, more attention should be given to the usage of these disinfectants, and their fate in nontarget environments should be monitored more tightly.

scholarly journals Intestinal Microbiota Containing Barnesiella Species Cures Vancomycin-Resistant Enterococcus faecium Colonization

2013 ◽  
Vol 81 (3) ◽  
pp. 965-973 ◽  
Author(s):  
Carles Ubeda ◽  
Vanni Bucci ◽  
Silvia Caballero ◽  
Ana Djukovic ◽  
Nora C. Toussaint ◽  
...  
Keyword(s):  
Intestinal Microbiota ◽  
Resistant Bacteria ◽  
Rrna Gene ◽  
Intestinal Colonization ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Hematopoietic Stem ◽  
Content Type ◽  
Obligate Anaerobic ◽  
Vancomycin Resistant

ABSTRACTBacteria causing infections in hospitalized patients are increasingly antibiotic resistant. Classical infection control practices are only partially effective at preventing spread of antibiotic-resistant bacteria within hospitals. Because the density of intestinal colonization by the highly antibiotic-resistant bacterium vancomycin-resistantEnterococcus(VRE) can exceed 109organisms per gram of feces, even optimally implemented hygiene protocols often fail. Decreasing the density of intestinal colonization, therefore, represents an important approach to limit VRE transmission. We demonstrate that reintroduction of a diverse intestinal microbiota to densely VRE-colonized mice eliminates VRE from the intestinal tract. While oxygen-tolerant members of the microbiota are ineffective at eliminating VRE, administration of obligate anaerobic commensal bacteria to mice results in a billionfold reduction in the density of intestinal VRE colonization. 16S rRNA gene sequence analysis of intestinal bacterial populations isolated from mice that cleared VRE following microbiota reconstitution revealed that recolonization with a microbiota that containsBarnesiellacorrelates with VRE elimination. Characterization of the fecal microbiota of patients undergoing allogeneic hematopoietic stem cell transplantation demonstrated that intestinal colonization withBarnesiellaconfers resistance to intestinal domination and bloodstream infection with VRE. Our studies indicate that obligate anaerobic bacteria belonging to theBarnesiellagenus enable clearance of intestinal VRE colonization and may provide novel approaches to prevent the spread of highly antibiotic-resistant bacteria.

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 Sourcing Antibiotic-Resistant Escherichia coli in Aquatic Ecosystems: A Combined Laboratory and Field Module

2021 ◽  
Author(s):  
Matthew J. Heard ◽  
Christopher E. Barton ◽  
Victoria J. Frost ◽  
Rachel Hongo
Keyword(s):  
Escherichia Coli ◽  
Aquatic Ecosystems ◽  
Indicator Bacteria ◽  
Resistant Bacteria ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Content Type ◽  
Laboratory Activity

The emergence of antibiotic-resistant bacteria represents a growing threat in aquatic ecosystems. In this combined field and laboratory activity, students will determine whether Escherichia coli , an indicator bacteria species commonly found in aquatic ecosystems, shows signs of resistance to common antibiotics.

scholarly journals High-Throughput, Signature-Tagged Mutagenic Approach To Identify Novel Virulence Factors of Yersinia pestis CO92 in a Mouse Model of Infection

2015 ◽  
Vol 83 (5) ◽  
pp. 2065-2081 ◽  
Author(s):  
Duraisamy Ponnusamy ◽  
Eric C. Fitts ◽  
Jian Sha ◽  
Tatiana E. Erova ◽  
Elena V. Kozlova ◽  
...  
Keyword(s):  
Mouse Model ◽  
Yersinia Pestis ◽  
Virulence Factors ◽  
High Throughput ◽  
Molecular Mechanisms ◽  
Sugar Transport ◽  
Type Vi Secretion System ◽  
Pneumonic Plague ◽  
Content Type ◽  
Genes Encoding

The identification of new virulence factors inYersinia pestisand understanding their molecular mechanisms during an infection process are necessary in designing a better vaccine or to formulate an appropriate therapeutic intervention. By using a high-throughput, signature-tagged mutagenic approach, we created 5,088 mutants ofY. pestisstrain CO92 and screened them in a mouse model of pneumonic plague at a dose equivalent to 5 50% lethal doses (LD50) of wild-type (WT) CO92. From this screen, we obtained 118 clones showing impairment in disseminating to the spleen, based on hybridization of input versus output DNA from mutant pools with 53 unique signature tags. In the subsequent screen, 20/118 mutants exhibited attenuation at 8 LD50when tested in a mouse model of bubonic plague, with infection by 10/20 of the aforementioned mutants resulting in 40% or higher survival rates at an infectious dose of 40 LD50. Upon sequencing, six of the attenuated mutants were found to carry interruptions in genes encoding hypothetical proteins or proteins with putative functions. Mutants with in-frame deletion mutations of two of the genes identified from the screen, namely,rbsA, which codes for a putative sugar transport system ATP-binding protein, andvasK, a component of the type VI secretion system, were also found to exhibit some attenuation at 11 or 12 LD50in a mouse model of pneumonic plague. Likewise, among the remaining 18 signature-tagged mutants, 9 were also attenuated (40 to 100%) at 12 LD50in a pneumonic plague mouse model. Previously, we found that deleting genes encoding Braun lipoprotein (Lpp) and acyltransferase (MsbB), the latter of which modifies lipopolysaccharide function, reduced the virulence ofY. pestisCO92 in mouse models of bubonic and pneumonic plague. Deletion ofrbsAandvasKgenes from either the Δlppsingle or the ΔlppΔmsbBdouble mutant augmented the attenuation to provide 90 to 100% survivability to mice in a pneumonic plague model at 20 to 50 LD50. The mice infected with the ΔlppΔmsbBΔrbsAtriple mutant at 50 LD50were 90% protected upon subsequent challenge with 12 LD50of WT CO92, suggesting that this mutant or others carrying combinational deletions of genes identified through our screen could potentially be further tested and developed into a live attenuated plague vaccine(s).

scholarly journals Repeated Isolation of Extended-Spectrum-β-Lactamase-Positive Escherichia coli Sequence Types 648 and 131 from Community Wastewater Indicates that Sewage Systems Are Important Sources of Emerging Clones of Antibiotic-Resistant Bacteria

2019 ◽  
Vol 63 (9) ◽  
Author(s):  
Erik Paulshus ◽  
Kaisa Thorell ◽  
Jessica Guzman-Otazo ◽  
Enrique Joffre ◽  
Patricia Colque ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Resistant Bacteria ◽  
Antibiotic Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Content Type ◽  
E Coli ◽  
Extended Spectrum ◽  
Pump Station ◽  
Sequence Types

ABSTRACT Antibiotic resistance in bacteria is an emerging problem globally. Resistant bacteria are found in human and animal microbiota, as well as in the environment. Wastewater receives bacteria from all these sources and thus can provide a measurement of abundance and diversity of antibiotic-resistant bacteria circulating in communities. In this study, water samples were collected from a wastewater pump station in a Norwegian suburban community over a period of 15 months. A total of 45 daily samples were cultured and analyzed for the presence of Escherichia coli. Eighty E. coli-like colonies were collected from each daily sample and then phenotyped and analyzed for antibiotic resistance using the PhenePlate-AREB system. During the sampling period, two unique E. coli phenotypes with resistance to cefotaxime and cefpodoxime indicating carriage of extended-spectrum β-lactamases (ESBL) were observed repeatedly. Whole-genome sequencing of 15 representative isolates from the two phenotypes identified these as two distinct clones belonging to the two globally spread E. coli multilocus sequence types (STs) ST131 and ST648 and carrying blaCTX-M-15. The number of ESBL-positive E. coli strains in the community wastewater pump station was 314 of 3,123 (10%) analyzed E. coli strains. Of the ESBL-positive isolates, 37% belonged to ST648, and 7% belonged to ST131. Repeated findings of CTX-M-15-positive ST648 and ST131 over time indicate that these STs are resident in the analyzed wastewater systems and/or circulate abundantly in the community.

Sign in / Sign up

Export Citation Format

Share Document