36 results on '"Swenson DL"'
Search Results
2. REINFORCEMENT CORROSION AND THE DURABILITY OF CONCRETE BRIDGES.
- Author
-
SWENSON, DL, LOTT, KM VINE-, GREGG, DJ, VINE-LOTT, KM, VASSIE, P, and ARSHAD, M
- Published
- 1985
- Full Text
- View/download PDF
3. Pneumonia in a long-term care facility. A prospective study of outcome.
- Author
-
Muder RR, Brennen C, Swenson DL, and Wagener M
- Published
- 1996
- Full Text
- View/download PDF
4. REINFORCEMENT CORROSION AND THE DURABILITY OF CONCRETE BRIDGES.
- Author
-
VASSIE, P, primary, SWENSON, DL, additional, GREGG, DJ, additional, VINE-LOTT, KM, additional, ARSHAD, M, additional, and LOTT, KM VINE-, additional
- Published
- 1985
- Full Text
- View/download PDF
5. Filovirus-like particles produced in insect cells: immunogenicity and protection in rodents.
- Author
-
Warfield KL, Posten NA, Swenson DL, Olinger GG, Esposito D, Gillette WK, Hopkins RF, Costantino J, Panchal RG, Hartley JL, Aman MJ, and Bavari S
- Abstract
BACKGROUND: Virus-like particles (VLPs) of Ebola virus (EBOV) and Marburg virus (MARV) produced in human 293T embryonic kidney cells have been shown to be effective vaccines against filoviral infection. In this study, we explored alternative strategies for production of filovirus-like particle-based vaccines, to accelerate the development process. The goal of this work was to increase the yield of VLPs, while retaining their immunogenic properties. METHODS: Ebola and Marburg VLPs (eVLPs and mVLPs, respectively) were generated by use of recombinant baculovirus constructs expressing glycoprotein, VP40 matrix protein, and nucleoprotein from coinfected insect cells. The baculovirus-derived eVLPs and mVLPs were characterized biochemically, and then the immune responses produced by the eVLPs in insect cells were studied further. RESULTS: The baculovirus-derived eVLPs elicited maturation of human myeloid dendritic cells (DCs), indicating their immunogenic properties. Mice vaccinated with insect cell-derived eVLPs generated antibody and cellular responses equivalent to those vaccinated with mammalian 293T cell-derived eVLPs and were protected from EBOV challenge in a dose-dependent manner. CONCLUSION: Together, these data suggest that filovirus-like particles produced by baculovirus expression systems, which are amenable to large-scale production, are highly immunogenic and are suitable as safe and effective vaccines for the prevention of filoviral infection. [ABSTRACT FROM AUTHOR]
- Published
- 2007
- Full Text
- View/download PDF
6. An Electronic Data Capture System and Nursing Research: An Integrative Health Intervention Design, Delivery, and Data Management Exemplar.
- Author
-
Withall J, Chau J, Coughlin V, Nash A, Grice-Swenson DL, Kaplan S, Marner V, Maydick-Youngberg D, Evanovich Zavotsky K, and Gabbe L
- Subjects
- Humans, Electronics, Data Management, Nursing Research
- Published
- 2024
- Full Text
- View/download PDF
7. Role of Antibodies in Protection Against Ebola Virus in Nonhuman Primates Immunized With Three Vaccine Platforms.
- Author
-
Warfield KL, Howell KA, Vu H, Geisbert J, Wong G, Shulenin S, Sproule S, Holtsberg FW, Leung DW, Amarasinghe GK, Swenson DL, Bavari S, Kobinger GP, Geisbert TW, and Aman MJ
- Subjects
- Animals, Antibodies, Viral blood, Ebola Vaccines immunology, Glycoproteins immunology, Hemorrhagic Fever, Ebola mortality, Hydrogen-Ion Concentration, Macaca fascicularis, Nucleoproteins immunology, Virion immunology, Antibodies, Viral immunology, Hemorrhagic Fever, Ebola prevention & control, Vaccination
- Abstract
Background: Several vaccine platforms have been successfully evaluated for prevention of Ebola virus (EBOV) disease (EVD) in nonhuman primates and humans. Despite remarkable efficacy by multiple vaccines, the immunological correlates of protection against EVD are incompletely understood., Methods: We systematically evaluated the antibody response to various EBOV proteins in 79 nonhuman primates vaccinated with various EBOV vaccine platforms. We evaluated the serum immunoglobulin (Ig)G titers against EBOV glycoprotein (GP), the ability of the vaccine-induced antibodies to bind GP at acidic pH or to displace ZMapp, and virus neutralization titers. The correlation of these outcomes with survival from EVD was evaluated by appropriate statistical methods., Results: Irrespective of the vaccine platform, protection from EVD strongly correlated with anti-GP IgG titers. The GP-directed antibody levels required for protection in animals vaccinated with virus-like particles (VLPs) lacking nucleoprotein (NP) was significantly higher than animals immunized with NP-containing VLPs or adenovirus-expressed GP, platforms that induce strong T-cell responses. Furthermore, protective immune responses correlated with anti-GP antibody binding strength at acidic pH, neutralization of GP-expressing pseudovirions, and the ability to displace ZMapp components from GP., Conclusions: These findings suggest key quantitative and qualitative attributes of antibody response to EVD vaccines as potential correlates of protection.
- Published
- 2018
- Full Text
- View/download PDF
8. AVI-7288 for Marburg Virus in Nonhuman Primates and Humans.
- Author
-
Heald AE, Charleston JS, Iversen PL, Warren TK, Saoud JB, Al-Ibrahim M, Wells J, Warfield KL, Swenson DL, Welch LS, Sazani P, Wong M, Berry D, Kaye EM, and Bavari S
- Subjects
- Animals, Antiviral Agents adverse effects, Antiviral Agents pharmacokinetics, Disease Models, Animal, Dose-Response Relationship, Drug, Humans, Kaplan-Meier Estimate, Macaca fascicularis, Marburg Virus Disease mortality, Morpholinos adverse effects, Morpholinos pharmacokinetics, RNA, Messenger, RNA, Viral, Antiviral Agents administration & dosage, Marburg Virus Disease drug therapy, Marburgvirus genetics, Morpholinos administration & dosage
- Abstract
Background: AVI-7288 is a phosphorodiamidate morpholino oligomer with positive charges that targets the viral messenger RNA that encodes Marburg virus (MARV) nucleoprotein. Its safety in humans is undetermined., Methods: We assessed the efficacy of AVI-7288 in a series of studies involving a lethal challenge with MARV in nonhuman primates. The safety of AVI-7288 was evaluated in a randomized, multiple-ascending-dose study in which 40 healthy humans (8 humans per dose group) received 14 once-daily infusions of AVI-7288 (1 mg, 4 mg, 8 mg, 12 mg, or 16 mg per kilogram of body weight) or placebo, in a 3:1 ratio. We estimated the protective dose in humans by comparing pharmacokinetic variables in infected nonhuman primates, uninfected nonhuman primates, and uninfected humans., Results: Survival in infected nonhuman primates was dose-dependent, with survival rates of 0%, 30%, 59%, 87%, 100%, and 100% among monkeys treated with 0 mg, 3.75 mg, 7.5 mg, 15 mg, 20 mg, and 30 mg of AVI-7288 per kilogram, respectively (P<0.001 with the use of the log-rank test for the comparison of survival across groups). No safety concern was identified at doses up to 16 mg per kilogram per day in humans. No serious adverse events were reported. Drug exposure (the area under the curve) was dose-dependent in both nonhuman primates and humans; drug clearance was independent of dose but was higher in nonhuman primates than in humans. The protective dose in humans was initially estimated, on the basis of exposure, to be 9.6 mg per kilogram per day (95% confidence interval, 6.6 to 12.5) for 14 days. Monte Carlo simulations supported a dose of 11 mg per kilogram per day to match the geometric mean protective exposure in nonhuman primates., Conclusions: This study shows that, on the basis of efficacy in nonhuman primates and pharmacokinetic data in humans, AVI-7288 has potential as postexposure prophylaxis for MARV infection in humans. (Funded by the Department of Defense; ClinicalTrials.gov number, NCT01566877.).
- Published
- 2015
- Full Text
- View/download PDF
9. Homologous and heterologous protection of nonhuman primates by Ebola and Sudan virus-like particles.
- Author
-
Warfield KL, Dye JM, Wells JB, Unfer RC, Holtsberg FW, Shulenin S, Vu H, Swenson DL, Bavari S, and Aman MJ
- Subjects
- Animals, Cross Protection immunology, Hemorrhagic Fever, Ebola blood, Hemorrhagic Fever, Ebola virology, Immunity, Humoral, Macaca blood, Macaca immunology, Macaca virology, Vaccination, Viral Vaccines immunology, Ebolavirus immunology, Hemorrhagic Fever, Ebola immunology, Hemorrhagic Fever, Ebola prevention & control, Virion immunology
- Abstract
Filoviruses cause hemorrhagic fever resulting in significant morbidity and mortality in humans. Several vaccine platforms that include multiple virus-vectored approaches and virus-like particles (VLPs) have shown efficacy in nonhuman primates. Previous studies have shown protection of cynomolgus macaques against homologous infection for Ebola virus (EBOV) and Marburg virus (MARV) following a three-dose vaccine regimen of EBOV or MARV VLPs, as well as heterologous protection against Ravn Virus (RAVV) following vaccination with MARV VLPs. The objectives of the current studies were to determine the minimum number of vaccine doses required for protection (using EBOV as the test system) and then demonstrate protection against Sudan virus (SUDV) and Taï Forest virus (TAFV). Using the EBOV nonhuman primate model, we show that one or two doses of VLP vaccine can confer protection from lethal infection. VLPs containing the SUDV glycoprotein, nucleoprotein and VP40 matrix protein provide complete protection against lethal SUDV infection in macaques. Finally, we demonstrate protective efficacy mediated by EBOV, but not SUDV, VLPs against TAFV; this is the first demonstration of complete cross-filovirus protection using a single component heterologous vaccine within the Ebolavirus genus. Along with our previous results, this observation provides strong evidence that it will be possible to develop and administer a broad-spectrum VLP-based vaccine that will protect against multiple filoviruses by combining only three EBOV, SUDV and MARV components.
- Published
- 2015
- Full Text
- View/download PDF
10. Using mortality risk scores for long-term prognosis of nursing home residents: caution is recommended.
- Author
-
Kruse RL, Parker Oliver D, Mehr DR, Petroski GF, Swenson DL, and Zweig SC
- Subjects
- Aged, Aged, 80 and over, Female, Geriatric Assessment, Humans, Logistic Models, Male, Prognosis, Prospective Studies, Risk Assessment, Time Factors, Mortality trends, Nursing Homes statistics & numerical data
- Abstract
Background: Determining prognosis for nursing home residents is important for care planning, but reliable prediction is difficult. We compared performance of four long-term mortality risk indices for nursing home residents-the Minimum Data Set Mortality Risk Index (MMRI), a recent revision to this index (MMRI-R), and the original and revised Flacker-Kiely models., Methods: We conducted a prospective cohort study in one 92-bed facility in Missouri. Participants were 130 residents who received a Minimum Data Set assessment from May through October, 2007. We collected the Minimum Data Set variables needed to calculate the mortality risk scores. We determined 6- and 12-month mortality for included residents. Using each mortality risk score as the sole independent predictor in logistic models predicting mortality, we determined discrimination (c-statistic) and calibration (Hosmer-Lemeshow goodness-of-fit statistic) for each model., Results: In our sample, discrimination was 0.59 for both the MMRI and the MMRI-R. Discrimination of the original Flacker-Kiely model was 0.69 for both 6 months and 1 year and 0.71 and 0.70, respectively, for the revised model. Model calibration was adequate for all models., Conclusions: Performance of four models that predict long-term mortality of nursing home residents was fair. In our population, the Flacker-Kiely models had similar and markedly better discrimination than either the MMRI or the MMRI-R.
- Published
- 2010
- Full Text
- View/download PDF
11. Advanced antisense therapies for postexposure protection against lethal filovirus infections.
- Author
-
Warren TK, Warfield KL, Wells J, Swenson DL, Donner KS, Van Tongeren SA, Garza NL, Dong L, Mourich DV, Crumley S, Nichols DK, Iversen PL, and Bavari S
- Subjects
- Animals, Ebolavirus isolation & purification, Filoviridae Infections immunology, Filoviridae Infections mortality, Hemorrhagic Fever, Ebola genetics, Hemorrhagic Fever, Ebola immunology, Hemorrhagic Fever, Ebola prevention & control, Humans, Macaca mulatta immunology, Marburg Virus Disease genetics, Marburg Virus Disease immunology, Marburg Virus Disease prevention & control, Marburgvirus isolation & purification, Primates, Safety, Filoviridae Infections genetics, Filoviridae Infections prevention & control, Vaccines, Synthetic therapeutic use, Viral Vaccines therapeutic use
- Abstract
Currently, no vaccines or therapeutics are licensed to counter Ebola or Marburg viruses, highly pathogenic filoviruses that are causative agents of viral hemorrhagic fever. Here we show that administration of positively charged phosphorodiamidate morpholino oligomers (PMOplus), delivered by various dosing strategies initiated 30-60 min after infection, protects>60% of rhesus monkeys against lethal Zaire Ebola virus (ZEBOV) and 100% of cynomolgus monkeys against Lake Victoria Marburg virus (MARV) infection. PMOplus may be useful for treating these and other highly pathogenic viruses in humans.
- Published
- 2010
- Full Text
- View/download PDF
12. Chemical modifications of antisense morpholino oligomers enhance their efficacy against Ebola virus infection.
- Author
-
Swenson DL, Warfield KL, Warren TK, Lovejoy C, Hassinger JN, Ruthel G, Blouch RE, Moulton HM, Weller DD, Iversen PL, and Bavari S
- Subjects
- Animals, Chlorocebus aethiops, Disease Models, Animal, Dose-Response Relationship, Drug, Ebolavirus genetics, Ebolavirus physiology, Female, Hemorrhagic Fever, Ebola virology, Humans, Male, Mice, Mice, Inbred C57BL, Rabbits, Structure-Activity Relationship, Treatment Outcome, Vero Cells, Virus Replication drug effects, Antiviral Agents chemistry, Antiviral Agents pharmacology, Antiviral Agents therapeutic use, Ebolavirus drug effects, Hemorrhagic Fever, Ebola drug therapy, Morpholines chemistry, Morpholines pharmacology, Morpholines therapeutic use, Oligonucleotides, Antisense chemistry, Oligonucleotides, Antisense pharmacology, Oligonucleotides, Antisense therapeutic use
- Abstract
Phosphorodiamidate morpholino oligomers (PMOs) are uncharged nucleic acid-like molecules designed to inactivate the expression of specific genes via the antisense-based steric hindrance of mRNA translation. PMOs have been successful at knocking out viral gene expression and replication in the case of acute viral infections in animal models and have been well tolerated in human clinical trials. We propose that antisense PMOs represent a promising class of therapeutic agents that may be useful for combating filoviral infections. We have previously shown that mice treated with a PMO whose sequence is complementary to a region spanning the start codon of VP24 mRNA were protected against lethal Ebola virus challenge. In the present study, we report on the abilities of two additional VP24-specific PMOs to reduce the cell-free translation of a VP24 reporter, to inhibit the in vitro replication of Ebola virus, and to protect mice against lethal challenge when the PMOs are delivered prior to infection. Additionally, structure-activity relationship evaluations were conducted to assess the enhancement of antiviral efficacy associated with PMO chemical modifications that included conjugation with peptides of various lengths and compositions, positioning of conjugated peptides to either the 5' or the 3' terminus, and the conferring of charge modifications by the addition of piperazine moieties. Conjugation with arginine-rich peptides greatly enhanced the antiviral efficacy of VP24-specific PMOs in infected cells and mice during lethal Ebola virus challenge.
- Published
- 2009
- Full Text
- View/download PDF
13. Monovalent virus-like particle vaccine protects guinea pigs and nonhuman primates against infection with multiple Marburg viruses.
- Author
-
Swenson DL, Warfield KL, Larsen T, Alves DA, Coberley SS, and Bavari S
- Subjects
- Amino Acid Sequence, Animal Structures pathology, Animal Structures virology, Animals, Antibodies, Viral blood, Antigens, Viral genetics, Guinea Pigs, Macaca fascicularis, Marburgvirus immunology, Molecular Sequence Data, Sequence Alignment, Survival Analysis, Vaccines, Inactivated immunology, Viral Plaque Assay, Viral Vaccines immunology, Viremia prevention & control, Antigens, Viral immunology, Marburg Virus Disease prevention & control, Marburgvirus genetics, Virosomes immunology
- Abstract
Background: Virus-like particle (VLP)-based vaccines have the advantage of being morphologically and antigenically similar to the live virus from which they are derived. Expression of the glycoprotein and VP40 matrix protein from Lake Victoria marburgvirus (MARV) results in spontaneous production of VLPs in mammalian cells. Guinea pigs vaccinated with Marburg virus VLPs (mVLPs) or inactivated MARV (iMARV) develop homologous humoral and T-cell responses and are completely protected from a lethal homologous MARV challenge., Aims & Methods: To determine whether mVLPs based on the Musoke (aka Lake Victoria) isolate of MARV could broadly protect against diverse isolates of MARV, guinea pigs were vaccinated with mVLPs or iMARV-Musoke and challenged with MARV-Musoke, -Ravn or -Ci67., Results: Prior to challenge, the mVLP- and iMARV-vaccinated guinea pigs had high levels of homologous MARV-Musoke and heterologous MARV-Ravn and -Ci67 antibodies. The Musoke-based mVLPs and iMARV vaccines provided complete protection in guinea pigs against viremia, viral replication and pathological changes in tissues, and lethal disease following challenge with MARV-Musoke, -Ravn or -Ci67. Guinea pigs vaccinated with RIBI adjuvant alone and infected with guinea pig-adapted MARV-Musoke, -Ravn or -Ci67 had histopathologic findings similar to those seen in the nonhuman primate model for MARV infection. Based on the strong protection observed in guinea pigs, we next vaccinated cynomolgus macaques with Musoke-based mVLPs and showed the VLP-vaccinated monkeys were broadly protected against three isolates of MARV (Musoke, Ravn and Ci67)., Conclusion: Musoke mVLPs are effective at inducing broad heterologous immunity and protection against multiple MARV isolates.
- Published
- 2008
- Full Text
- View/download PDF
14. Vaccine to confer to nonhuman primates complete protection against multistrain Ebola and Marburg virus infections.
- Author
-
Swenson DL, Wang D, Luo M, Warfield KL, Woraratanadharm J, Holman DH, Dong JY, and Pratt WD
- Subjects
- Adenoviridae immunology, Animals, Antigens, Viral genetics, Antigens, Viral metabolism, Bioterrorism prevention & control, Ebolavirus immunology, Ebolavirus pathogenicity, Hemorrhagic Fever, Ebola immunology, Humans, Macaca fascicularis, Marburg Virus Disease immunology, Marburgvirus classification, Marburgvirus immunology, Marburgvirus pathogenicity, Adenoviridae genetics, Ebola Vaccines administration & dosage, Ebola Vaccines genetics, Ebola Vaccines immunology, Filoviridae classification, Filoviridae genetics, Filoviridae immunology, Genetic Vectors, Hemorrhagic Fever, Ebola prevention & control, Marburg Virus Disease prevention & control, Viral Vaccines administration & dosage, Viral Vaccines genetics, Viral Vaccines immunology
- Abstract
Filoviruses (Ebola and Marburg viruses) are among the deadliest viruses known to mankind, with mortality rates nearing 90%. These pathogens are highly infectious through contact with infected body fluids and can be easily aerosolized. Additionally, there are currently no licensed vaccines available to prevent filovirus outbreaks. Their high mortality rates and infectious capabilities when aerosolized and the lack of licensed vaccines available to prevent such infectious make Ebola and Marburg viruses serious bioterrorism threats, placing them both on the category A list of bioterrorism agents. Here we describe a panfilovirus vaccine based on a complex adenovirus (CAdVax) technology that expresses multiple antigens from five different filoviruses de novo. Vaccination of nonhuman primates demonstrated 100% protection against infection by two species of Ebola virus and three Marburg virus subtypes, each administered at 1,000 times the lethal dose. This study indicates the feasibility of vaccination against all current filovirus threats in the event of natural hemorrhagic fever outbreak or biological attack.
- Published
- 2008
- Full Text
- View/download PDF
15. Ebola virus-like particle-based vaccine protects nonhuman primates against lethal Ebola virus challenge.
- Author
-
Warfield KL, Swenson DL, Olinger GG, Kalina WV, Aman MJ, and Bavari S
- Subjects
- Animals, Antibodies, Viral blood, Cell Line, Disease Models, Animal, Hemorrhagic Fever, Ebola blood, Humans, Kidney, Macaca fascicularis, Ebola Vaccines therapeutic use, Ebolavirus immunology, Hemorrhagic Fever, Ebola immunology
- Abstract
Background: Currently, there are no licensed vaccines or therapeutics for the prevention or treatment of infection by the highly lethal filoviruses, Ebola virus (EBOV) and Marburg virus (MARV), in humans. We previously had demonstrated the protective efficacy of virus-like particle (VLP)-based vaccines against EBOV and MARV infection in rodents., Methods: To determine the efficacy of vaccination with Ebola VLPs (eVLPs) in nonhuman primates, we vaccinated cynomolgus macaques with eVLPs containing EBOV glycoprotein (GP), nucleoprotein (NP), and VP40 matrix protein and challenged the macaques with 1000 pfu of EBOV., Results: Serum samples from the eVLP-vaccinated nonhuman primates demonstrated EBOV-specific antibody titers, as measured by enzyme-linked immunosorbent assay, complement-mediated lysis assay, and antibody-dependent cell-mediated cytotoxicity assay. CD44+ T cells from eVLP-vaccinated macaques but not from a naive macaque responded with vigorous production of tumor necrosis factor- alpha after EBOV-peptide stimulation. All 5 eVLP-vaccinated monkeys survived challenge without clinical or laboratory signs of EBOV infection, whereas the control animal died of infection., Conclusion: On the basis of safety and efficacy, eVLPs represent a promising filovirus vaccine for use in humans.
- Published
- 2007
- Full Text
- View/download PDF
16. Involvement of vacuolar protein sorting pathway in Ebola virus release independent of TSG101 interaction.
- Author
-
Silvestri LS, Ruthel G, Kallstrom G, Warfield KL, Swenson DL, Nelle T, Iversen PL, Bavari S, and Aman MJ
- Subjects
- Cell Line, Cell Membrane physiology, Cell Membrane virology, DNA Primers, Endosomal Sorting Complexes Required for Transport, Humans, Kidney, Microscopy, Confocal, Mutagenesis, Oligodeoxyribonucleotides, Protein Transport physiology, DNA-Binding Proteins physiology, Ebolavirus physiology, Transcription Factors physiology, Vacuoles physiology, Viral Proteins physiology
- Abstract
Budding of Ebola virus (EBOV) particles from the plasma membrane of infected cells requires viral and host proteins. EBOV virus matrix protein VP40 recruits TSG101, an ESCRT-1 (host cell endosomal sorting complex required for transport-1) complex protein in the vacuolar protein sorting (vps) pathway, to the plasma membrane during budding. Involvement of other vps proteins in EBOV budding has not been established. Therefore, we used VP40 deletion analysis, virus-like particle-release assays, and confocal microscopy to investigate the potential role of ESCRT-1 proteins VPS4, VPS28, and VPS37B in EBOV budding. We found that VP40 could redirect each protein from endosomes to the cell surface independently of TSG101 interaction. A lack of VPS4 adenosine triphosphatase activity reduced budding by up to 80%. Inhibition of VPS4 gene expression by use of phosphorodiamidite morpholino antisense oligonucleotides protected mice from lethal EBOV infection. These data show that EBOV can use vps proteins independently of TSG101 for budding and reveal VPS4 as a potential target for filovirus therapeutics.
- Published
- 2007
- Full Text
- View/download PDF
17. Ebola virus inactivation with preservation of antigenic and structural integrity by a photoinducible alkylating agent.
- Author
-
Warfield KL, Swenson DL, Olinger GG, Kalina WV, Viard M, Aitichou M, Chi X, Ibrahim S, Blumenthal R, Raviv Y, Bavari S, and Aman MJ
- Subjects
- Alkylating Agents radiation effects, Animals, Antiviral Agents pharmacology, Chlorocebus aethiops, Democratic Republic of the Congo, Disease Models, Animal, Ebolavirus pathogenicity, Female, Haplorhini, Mice, Mice, Inbred C57BL, Photochemistry, Vero Cells, Alkylating Agents pharmacology, Antigens, Viral analysis, Azides pharmacology, Ebolavirus drug effects, Ebolavirus ultrastructure, Hemorrhagic Fever, Ebola immunology, Viral Vaccines therapeutic use
- Abstract
Current methods for inactivating filoviruses are limited to high doses of irradiation or formalin treatment, which may cause structural perturbations that are reflected by poor immunogenicity. In this report, we describe a novel inactivation technique for Zaire Ebola virus (ZEBOV) that uses the photoinduced alkylating probe 1,5-iodonaphthylazide (INA). INA is incorporated into lipid bilayers and, when activated by ultraviolet irradiation, alkylates the proteins therein. INA treatment of ZEBOV resulted in the complete loss of infectivity in cells. Results of electron microscopy and virus-capture assays suggested the preservation of conformational surface epitopes. Challenge with 50,000 pfu of INA-inactivated, mouse-adapted ZEBOV did not cause disease or death in mice. A single vaccination with INA-inactivated ZEBOV (equivalent to 5 x 10(4) pfu) protected mice against lethal challenge with 1000 pfu of ZEBOV. INA-inactivated virus induced a protective response in 100% of mice when administered 3 days before challenge. Thus, INA may have significant potential for the development of vaccines and immunotherapeutics for filoviruses and other enveloped viruses.
- Published
- 2007
- Full Text
- View/download PDF
18. NKp30-dependent cytolysis of filovirus-infected human dendritic cells.
- Author
-
Fuller CL, Ruthel G, Warfield KL, Swenson DL, Bosio CM, Aman MJ, and Bavari S
- Subjects
- Cell Death genetics, Cell Death physiology, Cells, Cultured, Cytokines metabolism, Cytotoxicity, Immunologic immunology, Dendritic Cells immunology, Dendritic Cells virology, Fas Ligand Protein metabolism, Flow Cytometry, Humans, K562 Cells, Killer Cells, Natural immunology, Killer Cells, Natural metabolism, Membrane Glycoproteins metabolism, Mutation, Natural Cytotoxicity Triggering Receptor 1, Natural Cytotoxicity Triggering Receptor 2, Natural Cytotoxicity Triggering Receptor 3, Perforin, Pore Forming Cytotoxic Proteins metabolism, Receptors, Immunologic genetics, Receptors, Immunologic metabolism, Virion growth & development, Dendritic Cells metabolism, Filoviridae growth & development, Receptors, Immunologic physiology
- Abstract
Understanding how protective innate immune responses are generated is crucial to defeating highly lethal emerging pathogens. Accumulating evidence suggests that potent innate immune responses are tightly linked to control of Ebola and Marburg filoviral infections. Here, we report that unlike authentic or inactivated Ebola and Marburg, filovirus-derived virus-like particles directly activated human natural killer (NK) cells in vitro, evidenced by pro-inflammatory cytokine production and enhanced cytolysis of permissive target cells. Further, we observed perforin- and CD95L-mediated cytolysis of filovirus-infected human dendritic cells (DCs), primary targets of filovirus infection, by autologous NK cells. Gene expression knock-down studies directly linked NK cell lysis of infected DCs to upregulation of the natural cytotoxicity receptor, NKp30. These results are the first to propose a role for NK cells in the clearance of infected DCs and the potential involvement of NKp30-mediated cytolysis in control of viral infection in vivo. Further elucidation of the biology of NK cell activation, specifically natural cytotoxicity receptors like NKp30 and NKp46, promises to aid our understanding of microbial pathology.
- Published
- 2007
- Full Text
- View/download PDF
19. Activation of triggering receptor expressed on myeloid cells-1 on human neutrophils by marburg and ebola viruses.
- Author
-
Mohamadzadeh M, Coberley SS, Olinger GG, Kalina WV, Ruthel G, Fuller CL, Swenson DL, Pratt WD, Kuhns DB, and Schmaljohn AL
- Subjects
- Adaptor Proteins, Signal Transducing, Animals, Chlorocebus aethiops, Ebolavirus metabolism, Hemorrhagic Fever, Ebola immunology, Hemorrhagic Fever, Ebola metabolism, Hemorrhagic Fever, Ebola mortality, Humans, Immunity, Innate immunology, Inflammation immunology, Inflammation metabolism, Inflammation mortality, Marburg Virus Disease immunology, Marburg Virus Disease metabolism, Marburg Virus Disease mortality, Marburgvirus metabolism, Membrane Glycoproteins biosynthesis, Membrane Proteins, Phosphorylation, Protein Processing, Post-Translational immunology, Receptors, Immunologic biosynthesis, Receptors, Immunologic metabolism, Sepsis immunology, Sepsis metabolism, Sepsis mortality, Triggering Receptor Expressed on Myeloid Cells-1, Vero Cells, Ebolavirus immunology, Marburgvirus immunology, Membrane Glycoproteins immunology, Neutrophil Activation immunology, Neutrophils metabolism, Neutrophils virology, Receptors, Immunologic immunology
- Abstract
Marburg virus (MARV) and Ebola virus (EBOV), members of the viral family Filoviridae, cause fatal hemorrhagic fevers in humans and nonhuman primates. High viral burden is coincident with inadequate adaptive immune responses and robust inflammatory responses, and virus-mediated dysregulation of early host defenses has been proposed. Recently, a novel class of innate receptors called the triggering receptors expressed in myeloid cells (TREM) has been discovered and shown to play an important role in innate inflammatory responses and sepsis. Here, we report that MARV and EBOV activate TREM-1 on human neutrophils, resulting in DAP12 phosphorylation, TREM-1 shedding, mobilization of intracellular calcium, secretion of proinflammatory cytokines, and phenotypic changes. A peptide specific to TREM-1 diminished the release of tumor necrosis factor alpha by filovirus-activated human neutrophils in vitro, and a soluble recombinant TREM-1 competitively inhibited the loss of cell surface TREM-1 that otherwise occurred on neutrophils exposed to filoviruses. These data imply direct activation of TREM-1 by filoviruses and also indicate that neutrophils may play a prominent role in the immune and inflammatory responses to filovirus infections.
- Published
- 2006
- Full Text
- View/download PDF
20. VP35 knockdown inhibits Ebola virus amplification and protects against lethal infection in mice.
- Author
-
Enterlein S, Warfield KL, Swenson DL, Stein DA, Smith JL, Gamble CS, Kroeker AD, Iversen PL, Bavari S, and Mühlberger E
- Subjects
- Animals, Chlorocebus aethiops, Dose-Response Relationship, Drug, Ebolavirus genetics, Female, Hemorrhagic Fever, Ebola immunology, Kinetics, Mice, Mice, Inbred C57BL, Peptides chemistry, RNA, Viral genetics, Reverse Transcriptase Polymerase Chain Reaction, Vero Cells, Viral Regulatory and Accessory Proteins, Virus Replication drug effects, Virus Replication genetics, Ebolavirus drug effects, Hemorrhagic Fever, Ebola mortality, Hemorrhagic Fever, Ebola prevention & control, Morpholines pharmacology, Viral Proteins pharmacology
- Abstract
Phosphorodiamidate morpholino oligomers (PMO) are a class of uncharged single-stranded DNA analogs modified such that each subunit includes a phosphorodiamidate linkage and morpholine ring. PMO antisense agents have been reported to effectively interfere with the replication of several positive-strand RNA viruses in cell culture. The filoviruses, Marburg virus and Ebola virus (EBOV), are negative-strand RNA viruses that cause up to 90% lethality in human outbreaks. There is currently no commercially available vaccine or efficacious therapeutic for any filovirus. In this study, PMO conjugated to arginine-rich cell-penetrating peptide (P-PMO) and nonconjugated PMO were assayed for the ability to inhibit EBOV infection in cell culture and in a mouse model of lethal EBOV infection. A 22-mer P-PMO designed to base pair with the translation start site region of EBOV VP35 positive-sense RNA generated sequence-specific and time- and dose-dependent inhibition of EBOV amplification in cell culture. The same oligomer provided complete protection to mice when administered before or after an otherwise lethal infection of EBOV. A corresponding nonconjugated PMO, as well as nonconjugated truncated versions of 16 and 19 base residues, provided length-dependent protection to mice when administered prophylactically. Together, these data suggest that antisense PMO and P-PMO have the potential to control EBOV infection and are promising therapeutic candidates.
- Published
- 2006
- Full Text
- View/download PDF
21. Gene-specific countermeasures against Ebola virus based on antisense phosphorodiamidate morpholino oligomers.
- Author
-
Warfield KL, Swenson DL, Olinger GG, Nichols DK, Pratt WD, Blouch R, Stein DA, Aman MJ, Iversen PL, and Bavari S
- Subjects
- Animals, Ebolavirus genetics, Ebolavirus physiology, Female, Macaca mulatta, Male, Mice, Mice, Inbred C57BL, Morpholinos, Protein Biosynthesis drug effects, RNA, Viral antagonists & inhibitors, Viral Proteins antagonists & inhibitors, Viral Regulatory and Accessory Proteins, Virus Replication drug effects, Antisense Elements (Genetics) pharmacology, Ebolavirus drug effects, Hemorrhagic Fever, Ebola prevention & control, Morpholines pharmacology
- Abstract
The filoviruses Marburg virus and Ebola virus (EBOV) quickly outpace host immune responses and cause hemorrhagic fever, resulting in case fatality rates as high as 90% in humans and nearly 100% in nonhuman primates. The development of an effective therapeutic for EBOV is a daunting public health challenge and is hampered by a paucity of knowledge regarding filovirus pathogenesis. This report describes a successful strategy for interfering with EBOV infection using antisense phosphorodiamidate morpholino oligomers (PMOs). A combination of EBOV-specific PMOs targeting sequences of viral mRNAs for the viral proteins (VPs) VP24, VP35, and RNA polymerase L protected rodents in both pre- and post-exposure therapeutic regimens. In a prophylactic proof-of-principal trial, the PMOs also protected 75% of rhesus macaques from lethal EBOV infection. The work described here may contribute to development of designer, "druggable" countermeasures for filoviruses and other microbial pathogens.
- Published
- 2006
- Full Text
- View/download PDF
22. Induction of humoral and CD8+ T cell responses are required for protection against lethal Ebola virus infection.
- Author
-
Warfield KL, Olinger G, Deal EM, Swenson DL, Bailey M, Negley DL, Hart MK, and Bavari S
- Subjects
- Adjuvants, Immunologic administration & dosage, Amino Acid Sequence, Animals, Antibodies, Viral physiology, CD8-Positive T-Lymphocytes metabolism, Cells, Cultured, Epitopes, T-Lymphocyte immunology, Female, Hemorrhagic Fever, Ebola immunology, Interferon-gamma biosynthesis, Male, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Mice, Knockout, Molecular Sequence Data, Saponins administration & dosage, Saponins immunology, Vaccines, Subunit administration & dosage, Vaccines, Subunit immunology, Viral Envelope Proteins immunology, Viral Matrix Proteins immunology, Viral Vaccines administration & dosage, Viral Vaccines immunology, Virion immunology, Antibodies, Viral biosynthesis, CD8-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes virology, Ebolavirus immunology, Hemorrhagic Fever, Ebola mortality, Hemorrhagic Fever, Ebola prevention & control
- Abstract
Ebola virus (EBOV)-like particles (eVLP), composed of the EBOV glycoprotein and matrix viral protein (VP)40 with a lipid membrane, are a highly efficacious method of immunization against EBOV infection. The exact requirements for immunity against EBOV infection are poorly defined at this time. The goal of this work was to determine the requirements for EBOV immunity following eVLP vaccination. Vaccination of BALB/c or C57BL/6 mice with eVLPs in conjunction with QS-21 adjuvant resulted in mixed IgG subclass responses, a Th1-like memory cytokine response, and protection from lethal EBOV challenge. Further, this vaccination schedule led to the generation of both CD4(+) and CD8(+) IFN-gamma(+) T cells recognizing specific peptides within glycoprotein and VP40. The transfer of both serum and splenocytes, but not serum or splenocytes alone, from eVLP-vaccinated mice conferred protection against lethal EBOV infection in these studies. B cells were required for eVLP-mediated immunity to EBOV because B cell-deficient mice vaccinated with eVLPs were not protected from lethal EBOV challenge. We also found that CD8(+), but not CD4(+), T cells are absolutely required for eVLP-mediated protection against EBOV infection. Further, eVLP-induced protective mechanisms were perforin-independent, but IFN-gamma-dependent. Taken together, both EBOV-specific humoral and cytotoxic CD8(+) T cell responses are critical to mediate protection against filoviruses following eVLP vaccination.
- Published
- 2005
- Full Text
- View/download PDF
23. Analysis of Ebola virus and VLP release using an immunocapture assay.
- Author
-
Kallstrom G, Warfield KL, Swenson DL, Mort S, Panchal RG, Ruthel G, Bavari S, and Aman MJ
- Subjects
- Animals, Cell Line, Ebolavirus genetics, Nucleocapsid Proteins analysis, Nucleoproteins, Transfection, Viral Core Proteins, Ebolavirus isolation & purification, Enzyme-Linked Immunosorbent Assay methods
- Abstract
Ebola virus (EBOV), an emerging pathogen, is the causative agent of a rapidly progressive hemorrhagic fever with high mortality rates. There are currently no approved vaccines or treatments available for Ebola hemorrhagic fever. Standard plaque assays are currently the only reliable techniques for enumerating the virus. Effective drug-discovery screening as well as target identification and validation require simple and more rapid detection methods. This report describes the development of a rapid ELISA that measures virus release with high sensitivity. This assay detects both Ebola virus and EBOV-like particles (VLPs) directly from cell-culture supernatants with the VP40 matrix protein serving as antigen. Using this assay, the contribution of the EBOV nucleocapsid (NC) proteins in VLP release was determined. These findings indicate that a combination of NC proteins together with the envelope components is optimal for VLP formation and release, a finding that is important for vaccination with Ebola VLPs. Furthermore, this assay can be used in surrogate models in non-biocontainment environment, facilitating both basic research on the mechanism of EBOV assembly and budding as well as drug-discovery research.
- Published
- 2005
- Full Text
- View/download PDF
24. Filovirus-like particles as vaccines and discovery tools.
- Author
-
Warfield KL, Swenson DL, Demmin G, and Bavari S
- Subjects
- Animals, Filoviridae Infections virology, Humans, Technology, Pharmaceutical methods, Filoviridae, Filoviridae Infections prevention & control, Technology, Pharmaceutical trends, Viral Vaccines administration & dosage
- Abstract
Ebola and Marburg viruses are members of the family Filoviridae, which cause severe hemorrhagic fevers in humans. Filovirus outbreaks have been sporadic, with mortality rates currently ranging from 30 to 90%. Unfortunately, there is no efficacious human therapy or vaccine available to treat disease caused by either Ebola or Marburg virus infection. Expression of the filovirus matrix protein, VP40, is sufficient to drive spontaneous production and release of virus-like particles (VLPs) that resemble the distinctively filamentous infectious virions. The addition of other filovirus proteins, including virion proteins (VP)24, 30 and 35 and glycoprotein, increases the efficiency of VLP production and results in particles containing multiple filovirus antigens. Vaccination with Ebola or Marburg VLPs containing glycoprotein and VP40 completely protects rodents from lethal challenge with the homologous virus. These candidate vaccines are currently being tested for immunogenicity and efficacy in nonhuman primates. Furthermore, the Ebola and Marburg VLPs are being used as a surrogate model to further understand the filovirus life cycle, with the goal of developing rationally designed vaccines and therapeutics. Thus, in addition to their use as a vaccine, VLPs are currently being used as tools to learn lessons about filovirus pathogenesis, immunology, replication and assembly requirements.
- Published
- 2005
- Full Text
- View/download PDF
25. Virus-like particles exhibit potential as a pan-filovirus vaccine for both Ebola and Marburg viral infections.
- Author
-
Swenson DL, Warfield KL, Negley DL, Schmaljohn A, Aman MJ, and Bavari S
- Subjects
- Animals, Antibodies, Viral blood, Glycoproteins immunology, Guinea Pigs, Nucleoproteins immunology, Vaccination, Viral Core Proteins immunology, Viremia prevention & control, Ebolavirus immunology, Marburgvirus immunology, Viral Vaccines immunology, Virion immunology
- Abstract
A safe and effective pan-filovirus vaccine is highly desirable since the filoviruses Ebola virus (EBOV) and Marburg virus (MARV) cause highly lethal disease typified by unimpeded viral replication and severe hemorrhagic fever. Previously, we showed that expression of the homologous glycoprotein (GP) and matrix protein VP40 from a single filovirus, either EBOV or MARV, resulted in formation of wild-type virus-like particles (VLPs) in mammalian cells. When used as a vaccine, the wild-type VLPs protected from homologous filovirus challenge. The aim of this work was to generate a multi-agent vaccine that would simultaneously protect against multiple and diverse members of the Filoviridae family. Our initial approach was to construct hybrid VLPs containing heterologous viral proteins, of EBOV and MARV, and test the efficacy of the hybrid VLPs in a guinea pig model. Our data indicate that vaccination with GP was required and sufficient to protect against a homologous filovirus challenge, as heterologous wild-type VLPs or hybrid VLPs that did not contain the homologous GP failed to protect. Alternately, we vaccinated guinea pigs with a mixture of wild-type Ebola and Marburg VLPs. Vaccination with a single dose of the multivalent VLP vaccine elicited strong immune responses to both viruses and protected animals against EBOV and MARV challenge. This work provides a critical foundation towards the development of a pan-filovirus vaccine that is safe and effective for use in primates and humans.
- Published
- 2005
- Full Text
- View/download PDF
26. Isolation of Mycobacterium tuberculosis mutants defective in the arrest of phagosome maturation.
- Author
-
Pethe K, Swenson DL, Alonso S, Anderson J, Wang C, and Russell DG
- Subjects
- Colloids chemistry, Genotype, Gold chemistry, Gold pharmacology, Hydrogen-Ion Concentration, Macrophages cytology, Macrophages microbiology, Macrophages ultrastructure, Microscopy, Electron, Mycobacterium tuberculosis isolation & purification, Mycobacterium tuberculosis ultrastructure, Phagosomes genetics, Phagosomes ultrastructure, Phenotype, Spectrometry, Fluorescence, Mutation genetics, Mycobacterium tuberculosis cytology, Mycobacterium tuberculosis genetics, Phagosomes physiology
- Abstract
Mycobacterium tuberculosis resides within the phagocytes of its host. It ensures its continued survival through arresting the normal maturation of its phagosome, which is retained within the early endosomal system of the macrophage. Although individual bacterial components have been shown to modulate phagosome biogenesis, the mechanism(s) active in live, intact bacteria remain elusive. We have developed a genetic screen that facilitates the isolation of mutants defective in arresting the maturation of their phagosomes. Macrophages were incubated with iron-dextran that was chased into lysosomes. The cells were subsequently infected with M. tuberculosis from a library of transposon-mutagenized bacteria. After four rounds of enrichment, the majority of mutants isolated were unable to prevent acidification of their phagosomes and were attenuated for intracellular survival. The genes affected range in function from those with no known homologues to putative transporters and lipid synthesis enzymes. Further characterization of these bacteria is needed. In addition to clarifying the processes active in modulation of phagosome biogenesis by M. tuberculosis, this screen may be applicable to other pathogens that restrict the maturation of their phagosome.
- Published
- 2004
- Full Text
- View/download PDF
27. Marburg virus-like particles protect guinea pigs from lethal Marburg virus infection.
- Author
-
Warfield KL, Swenson DL, Negley DL, Schmaljohn AL, Aman MJ, and Bavari S
- Subjects
- Animals, Antibodies, Viral analysis, Antibodies, Viral biosynthesis, Antibody Formation immunology, CD4-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes immunology, Ebolavirus immunology, Ebolavirus ultrastructure, Guinea Pigs, Lymphocyte Count, Marburgvirus ultrastructure, Microscopy, Electron, Neutralization Tests, Vaccination, Vaccines, Attenuated immunology, Vaccines, Attenuated therapeutic use, Viral Plaque Assay, Viral Vaccines immunology, Marburg Virus Disease prevention & control, Marburgvirus immunology, Viral Vaccines therapeutic use
- Abstract
Ongoing outbreaks of filoviruses in Africa and concerns about their use in bioterrorism attacks have led to intense efforts to find safe and effective vaccines to prevent the high mortality associated with these viruses. We previously reported the generation of virus-like particles (VLPs) for the filoviruses, Marburg (MARV) and Ebola (EBOV) virus, and that vaccinating mice with Ebola VLPs (eVLPs) results in complete survival from a lethal EBOV challenge. The objective of this study was to determine the efficacy of Marburg VLPs (mVLPs) as a potential vaccine against lethal MARV infection in a guinea pig model. Guinea pigs vaccinated with mVLPs or inactivated MARV developed MARV-specific antibody titers, as tested by ELISA or plaque-reduction and neutralization assays and were completely protected from a MARV challenge over 2000 LD50. While eVLP vaccination induced high EBOV-specific antibody responses, it did not cross-protect against MARV challenge in guinea pigs. Vaccination with mVLP or eVLP induced proliferative responses in vitro only upon re-exposure to the homologous antigen and this recall proliferative response was dependent on the presence of CD4+ T cells. Taken together with our previous work, these findings suggest that VLPs are a promising vaccine candidate for the deadly filovirus infections.
- Published
- 2004
- Full Text
- View/download PDF
28. Role of natural killer cells in innate protection against lethal ebola virus infection.
- Author
-
Warfield KL, Perkins JG, Swenson DL, Deal EM, Bosio CM, Aman MJ, Yokoyama WM, Young HA, and Bavari S
- Subjects
- Animals, Cytokines biosynthesis, Cytotoxicity, Immunologic, Humans, Immunotherapy methods, Interferon-gamma metabolism, Interleukin-2 metabolism, Membrane Glycoproteins metabolism, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Microscopy, Atomic Force, Perforin, Pore Forming Cytotoxic Proteins, Spleen metabolism, Time Factors, Hemorrhagic Fever, Ebola immunology, Hemorrhagic Fever, Ebola prevention & control, Killer Cells, Natural metabolism
- Abstract
Ebola virus is a highly lethal human pathogen and is rapidly driving many wild primate populations toward extinction. Several lines of evidence suggest that innate, nonspecific host factors are potentially critical for survival after Ebola virus infection. Here, we show that nonreplicating Ebola virus-like particles (VLPs), containing the glycoprotein (GP) and matrix protein virus protein (VP)40, administered 1-3 d before Ebola virus infection rapidly induced protective immunity. VLP injection enhanced the numbers of natural killer (NK) cells in lymphoid tissues. In contrast to live Ebola virus, VLP treatment of NK cells enhanced cytokine secretion and cytolytic activity against NK-sensitive targets. Unlike wild-type mice, treatment of NK-deficient or -depleted mice with VLPs had no protective effect against Ebola virus infection and NK cells treated with VLPs protected against Ebola virus infection when adoptively transferred to naive mice. The mechanism of NK cell-mediated protection clearly depended on perforin, but not interferon-gamma secretion. Particles containing only VP40 were sufficient to induce NK cell responses and provide protection from infection in the absence of the viral GP. These findings revealed a decisive role for NK cells during lethal Ebola virus infection. This work should open new doors for better understanding of Ebola virus pathogenesis and direct the development of immunotherapeutics, which target the innate immune system, for treatment of Ebola virus infection.
- Published
- 2004
- Full Text
- View/download PDF
29. Generation of Marburg virus-like particles by co-expression of glycoprotein and matrix protein.
- Author
-
Swenson DL, Warfield KL, Kuehl K, Larsen T, Hevey MC, Schmaljohn A, Bavari S, and Aman MJ
- Subjects
- Animals, Antibodies, Viral blood, Cell Line, Cells, Cultured, Female, Immunohistochemistry, Marburgvirus immunology, Membrane Glycoproteins metabolism, Mice, Mice, Inbred BALB C, Microscopy, Electron, Transfection, Viral Envelope Proteins genetics, Viral Envelope Proteins immunology, Viral Matrix Proteins genetics, Viral Matrix Proteins immunology, Virion immunology, Marburgvirus physiology, Viral Envelope Proteins metabolism, Viral Matrix Proteins metabolism, Virion metabolism
- Abstract
Marburg virus (MARV), the causative agent of a severe hemorrhagic fever, has a characteristic filamentous morphology. Here we report that co-expression of MARV glycoprotein and matrix protein (VP40) in mammalian cells leads to spontaneous budding of filamentous particles strikingly similar to wild-type MARV. In addition, these particles elicit an immune response in BALB/c mice. The generation of non-replicating Marburg virus-like particles (VLPs) should significantly facilitate the research on molecular mechanisms of MARV assembly and release. Furthermore, VLPs may be an excellent vaccine candidate against Marburg infection.
- Published
- 2004
- Full Text
- View/download PDF
30. Characterization of activity and expression of isocitrate lyase in Mycobacterium avium and Mycobacterium tuberculosis.
- Author
-
Höner Zu Bentrup K, Miczak A, Swenson DL, and Russell DG
- Subjects
- Acetates metabolism, Cloning, Molecular, Escherichia coli metabolism, Gene Expression Regulation, Enzymologic, Hydrogen-Ion Concentration, Isocitrate Lyase chemistry, Isocitrate Lyase isolation & purification, Isoenzymes chemistry, Isoenzymes isolation & purification, Isoenzymes metabolism, Kinetics, Open Reading Frames, Palmitates metabolism, Recombinant Proteins metabolism, Succinic Acid metabolism, Bacterial Proteins, Isocitrate Lyase metabolism, Mycobacterium avium enzymology, Mycobacterium tuberculosis enzymology
- Abstract
Analysis by two-dimensional gel electrophoresis revealed that Mycobacterium avium expresses several proteins unique to an intracellular infection. One abundant protein with an apparent molecular mass of 50 kDa was isolated, and the N-terminal sequence was determined. It matches a sequence in the M. tuberculosis database (Sanger) with similarity to the enzyme isocitrate lyase of both Corynebacterium glutamicum and Rhodococcus fascians. Only marginal similarity was observed between this open reading frame (ORF) (termed icl) and a second distinct ORF (named aceA) which exhibits a low similarity to other isocitrate lyases. Both ORFs can be found as distinct genes in the various mycobacterial databases recently published. Isocitrate lyase is a key enzyme in the glyoxylate cycle and is essential as an anapleurotic enzyme for growth on acetate and certain fatty acids as carbon source. In this study we express and purify Icl, as well as AceA proteins, and show that both exhibit isocitrate lyase activity. Various known inhibitors for isocitrate lyase were effective. Furthermore, we present evidence that in both M. avium and M. tuberculosis the production and activity of the isocitrate lyase is enhanced under minimal growth conditions when supplemented with acetate or palmitate.
- Published
- 1999
- Full Text
- View/download PDF
31. Two pathogenicity islands in uropathogenic Escherichia coli J96: cosmid cloning and sample sequencing.
- Author
-
Swenson DL, Bukanov NO, Berg DE, and Welch RA
- Subjects
- Base Sequence, Cloning, Molecular, Cosmids, DNA, Bacterial genetics, Escherichia coli genetics, Molecular Sequence Data, Restriction Mapping, Sequence Alignment, Sequence Homology, Nucleic Acid, Escherichia coli pathogenicity, Genes, Bacterial
- Abstract
Many of the virulence genes of pathogenic strains of Escherichia coli are carried in large multigene chromosomal segments called pathogenicity islands (PAIs) that are absent from normal fecal and laboratory K-12 strains of this bacterium. We are studying PAIs in order to better understand factors that govern virulence and to assess how such DNA segments are gained or lost during evolution. The isolation and sample sequencing of a set of 11 cosmid clones that cover all of one and much of a second large PAI in the uropathogenic E. coli J96 are described. These PAIs were mapped to the 64- and 94-min regions of the E. coli K-12 chromosome, which differ from the locations of three PAIs identified in other pathogenic E. coli strains. Analysis of the junction sequences with E. coli K-12-like DNAs showed that the insert at 94 min is within the 3' end of a phenylalanine tRNA gene, pheR, and is flanked by a 135-bp imperfect direct repeat. Analysis of the one junction recovered from the insert at 64 min indicated that it lies near another tRNA gene, pheV. To identify possible genes unique to these PAIs, 100 independent subclones of the cosmids were made by PstI digestion and ligation into a pBS+ plasmid and used in one-pass sample DNA sequencing from primer binding sites at the cloning site in the vector DNA. Database searches of the J96 PAI-specific sequences identified numerous instances in which the cloned DNAs shared significant sequence similarities to adhesins, toxins, and other virulence determinants of diverse pathogens. Several likely insertion sequence elements (IS100, IS630, and IS911) and conjugative R1 plasmid and P4 phage genes were also found. We propose that such mobile genetic elements may have facilitated the spread of virulence determinants within PAIs among bacteria.
- Published
- 1996
- Full Text
- View/download PDF
32. Battling against host phagocytes: the wherefore of the RTX family of toxins?
- Author
-
Welch RA, Bauer ME, Kent AD, Leeds JA, Moayeri M, Regassa LB, and Swenson DL
- Subjects
- Antitoxins immunology, Bacterial Proteins classification, Bacterial Proteins genetics, Bacterial Proteins metabolism, Bacterial Toxins classification, Bacterial Toxins genetics, Hemolysin Proteins, Phagocytes drug effects, Phagocytes metabolism, Bacterial Toxins metabolism
- Abstract
The RTX family of bacterial exotoxins is a group of related cytolytic proteins produced by a wide variety of gram-negative human and animal pathogens. While diverse in their associated diseases and in their target cell specificities, there remain several themes common to RTX toxins, including genetic organization, structural and functional features, and effects on target cells. In this review, we summarize and discuss the genetics, regulation, epidemiology, structure/function relationships, and in vivo and in vitro activities of the best characterized RTX toxins, and speculate on their roles in pathogenesis and their use in immunotherapy.
- Published
- 1995
33. The gene fimU affects expression of Salmonella typhimurium type 1 fimbriae and is related to the Escherichia coli tRNA gene argU.
- Author
-
Swenson DL, Kim KJ, Six EW, and Clegg S
- Subjects
- Agglutination, Bacteriophage lambda growth & development, Lysogeny, Restriction Mapping, Salmonella typhimurium growth & development, Salmonella typhimurium physiology, Escherichia coli genetics, Fimbriae, Bacterial physiology, Gene Expression Regulation, Bacterial, Genes, Bacterial, Multigene Family, Salmonella Phages growth & development, Salmonella typhimurium genetics
- Abstract
The gene fimU, located on a recombinant plasmid carrying the Salmonella typhimurium type 1 fimbrial gene cluster is closely related to the Escherichia coli tRNA gene argU. The fimU gene complements an E. coli argU mutant that is a P2 lysogen, thereby allowing the phage P4 to grow in this strain but preventing the growth of phage lambda. In addition, fimU was shown to be involved in fimbrial expression since transformants of the E. coli argU mutant could produce fimbriae only in the presence of fimU but not in its absence, whereas in an E. coli argU+ strain fimbriation did not require the fimU gene.
- Published
- 1994
- Full Text
- View/download PDF
34. Identification of ancillary fim genes affecting fimA expression in Salmonella typhimurium.
- Author
-
Swenson DL and Clegg S
- Subjects
- Amino Acid Sequence, Bacterial Proteins biosynthesis, Base Sequence, DNA Mutational Analysis, DNA-Binding Proteins, Membrane Proteins, Molecular Sequence Data, RNA, Messenger biosynthesis, Recombinant Fusion Proteins, Transformation, Genetic, beta-Galactosidase biosynthesis, beta-Galactosidase genetics, Bacterial Proteins genetics, Fimbriae Proteins, Fimbriae, Bacterial physiology, Gene Expression Regulation, Bacterial, Genes, Bacterial genetics, Salmonella typhimurium genetics
- Abstract
Regulation of the gene, fimA, encoding the major fimbrial subunit of S. typhimurium S6704 was examined by using a lambda fimA-lacZ lysogen. Transformation of the lambda fimA-lacZ lysogen with various derivatives of the recombinant plasmid that encodes type 1 fimbrial expression, pISF101, indicated that two regions of this plasmid alter beta-galactosidase production. One plasmid is a deletion resulting in the loss of a 28-kDa polypeptide downstream of fimA, while the other plasmid encodes a 24- and a 27-kDa polypeptide. Northern (RNA) blot analyses indicated that the steady-state fimA mRNA levels of these transformants were high. In addition, phenotypic expression of type 1 fimbriae by agar-grown cultures is observed only in those transformants bearing plasmids which show increased beta-galactosidase and fimA mRNA levels.
- Published
- 1992
- Full Text
- View/download PDF
35. The frequency of fim genes among Salmonella serovars.
- Author
-
Swenson DL, Clegg S, and Old DC
- Subjects
- Blotting, Southern, Chromosome Mapping, DNA Probes, Escherichia coli genetics, Gene Expression Regulation, Bacterial, Genetic Variation, Salmonella typhimurium genetics, Serotyping, Transformation, Genetic, Adhesins, Bacterial, Bacterial Adhesion genetics, Bacterial Proteins genetics, Fimbriae, Bacterial physiology, Hemagglutinins genetics, Salmonella genetics
- Abstract
Salmonella serovars were examined for the presence of fim gene sequences using specific DNA probes. All strains, regardless of their ability to express surface-associated fimbriae, retain a considerable amount of DNA homologous to the gene probes used. The phenotypically nonfimbriae FIRN and non-FIRN strains of S. typhimurium retain detectable amounts of fim gene sequences and, therefore, may not be genotypically non-fimbriate. The MS adhesin can be expressed by type 2 fimbriate bacteria when they are transformed with discrete regions of the fim gene cluster. However, this conversion to a hemagglutinating phenotype is not associated with a small region of DNA. Therefore, the inability of type 2 fimbria-producing strains of Salmonella to mediate hemagglutination does not appear to be due to a small deletion in a single fim gene.
- Published
- 1991
- Full Text
- View/download PDF
36. Expression of type 1 fimbriae and mannose-sensitive hemagglutinin by recombinant plasmids.
- Author
-
Gerlach GF, Clegg S, Ness NJ, Swenson DL, Allen BL, and Nichols WA
- Subjects
- Enterobacteriaceae physiology, Enterobacteriaceae ultrastructure, Genetic Complementation Test, Mannose, Microscopy, Electron, Nucleic Acid Hybridization, Restriction Mapping, Sequence Homology, Nucleic Acid, Bacterial Adhesion, Enterobacteriaceae genetics, Fimbriae, Bacterial ultrastructure, Hemagglutinins genetics, Plasmids
- Abstract
Deletions within the cloned genes (fimA) encoding the type 1 major fimbrial subunits of two isolates of Klebsiella pneumoniae resulted in a nonfimbriate but hemagglutinating phenotype after transformation of Escherichia coli HB101 or ORN103. Phenotypic expression of type 1 fimbriae could be restored by transformation with plasmids containing the fimA genes of the fimbrial gene clusters from different strains. The surface fimbriae expressed were serologically identical to those of the polymerized product of the introduced fimA gene. The fimA gene products of Salmonella typhimurium and Serratia marcescens could utilize the accessory fimbrial genes of K. pneumoniae to produce surface-associated, hemagglutinating fimbriae. The relatedness of the type 1 fimbrial gene clusters from multiple isolates of members of the family Enterobacteriaceae was examined by DNA hybridization techniques. These analyses demonstrated little nucleotide sequence agreement among distinct genera of the enteric bacteria.
- Published
- 1989
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.