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  • American Society for Microbiology  (7)
  • 1
    Online Resource
    Online Resource
    Production and Preclinical Evaluation of Plasmodium falciparum MSP-1 19 and MSP-3 11 Chimeric Protein, PfMSP-Fu 24
    American Society for Microbiology ; 2014
    In:  Clinical and Vaccine Immunology Vol. 21, No. 6 ( 2014-06), p. 886-897
    Details
    In: Clinical and Vaccine Immunology, American Society for Microbiology, Vol. 21, No. 6 ( 2014-06), p. 886-897
    Abstract: A Plasmodium falciparum chimeric protein, PfMSP-Fu 24 , was constructed by genetically coupling immunodominant, conserved regions of two merozoite surface proteins, the 19-kDa region C-terminal region of merozoite surface protein 1 (PfMSP-1 19 ) and an 11-kDa conserved region of merozoite surface protein 3 (PfMSP-3 11 ), to augment the immunogenicity potential of these blood-stage malaria vaccine candidates. Here we describe an improved, efficient, and scalable process to produce high-quality PfMSP-Fu 24 . The chimeric protein was produced in Escherichia coli SHuffle T7 Express lysY cells that express disulfide isomerase DsbC. A two-step purification process comprising metal affinity followed by cation exchange chromatography was developed, and we were able to obtain PfMSP-Fu 24 with purity above 99% and with a considerable yield of 23 mg/liter. Immunogenicity of PfMSP-Fu 24 formulated with several adjuvants, including Adjuplex, Alhydrogel, Adjuphos, Alhydrogel plus glucopyranosyl lipid adjuvant, aqueous (GLA-AF), Adjuphos+GLA-AF, glucopyranosyl lipid adjuvant-stable emulsion (GLA-SE), and Freund's adjuvant, was evaluated. PfMSP-Fu 24 formulated with GLA-SE and Freund's adjuvant in mice and with Alhydrogel and Freund's adjuvant in rabbits produced high titers of PfMSP-1 19 and PfMSP-3 11 -specific functional antibodies. Some of the adjuvant formulations induced inhibitory antibody responses and inhibited in vitro growth of P. falciparum parasites in the presence as well as in the absence of human monocytes. These results suggest that PfMSP-Fu 24 can form a constituent of a multistage malaria vaccine.
    Type of Medium: Online Resource
    ISSN: 1556-6811 , 1556-679X
    URL: Article
    DOI: 10.1128/CVI.00179-14
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2014
    detail.hit.zdb_id: 1496863-0
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  • 2
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    Plasmodium falciparum Cysteine-Rich Protective Antigen (CyRPA) Elicits Detectable Levels of Invasion-Inhibitory Antibodies during Natural Infection in Humans
    American Society for Microbiology ; 2022
    In:  Infection and Immunity Vol. 90, No. 1 ( 2022-01-25)
    Details
    In: Infection and Immunity, American Society for Microbiology, Vol. 90, No. 1 ( 2022-01-25)
    Abstract: Plasmodium falciparum cysteine-rich protective antigen (CyRPA) is a conserved component of an essential erythrocyte invasion complex (RH5/Ripr/CyRPA) and a target of potent cross-strain parasite-neutralizing antibodies. While naturally acquired human RH5 antibodies have been functionally characterized, there are no similar reports on CyRPA. Thus, we analyzed the parasite-neutralizing activity of naturally acquired human CyRPA antibodies. In this regard, CyRPA human antibodies were measured and purified from malaria-infected plasma obtained from patients in central India and analyzed for their parasite neutralizing activity via in vitro growth inhibition assays (GIA). We report that, despite being susceptible to antibodies, CyRPA is a highly conserved antigen that does not appear to be under substantial immune selection pressure, as a very low acquisition rate for anti-CyRPA antibodies was reported in malaria-exposed Indians. We demonstrate for the first time that the small amounts of natural CyRPA antibodies exhibited functional parasite-neutralizing activity and that a CyRPA-based vaccine formulation induces highly potent antibodies in rabbits. Importantly, the vaccine-induced CyRPA antibodies exhibited a robust 50% inhibitory concentration (IC 50 ) of 21.96 μg/ml, which is comparable to the IC 50 of antibodies against the leading blood-stage vaccine candidate, reticulocyte-binding-like homologous protein 5 (RH5). Our data support CyRPA as a unique vaccine target that is highly susceptible to immune attack but is highly conserved compared to other leading candidates such as MSP-1 and AMA-1, further substantiating its promise as a leading blood-stage vaccine candidate.
    Type of Medium: Online Resource
    ISSN: 0019-9567 , 1098-5522
    URL: Article
    DOI: 10.1128/IAI.00377-21
    RVK:
    XA 10000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 1483247-1
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  • 3
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    Identification of a Potent Combination of Key Plasmodium falciparum Merozoite Antigens That Elicit Strain-Transcending Parasite-Neutralizing Antibodies
    American Society for Microbiology ; 2013
    In:  Infection and Immunity Vol. 81, No. 2 ( 2013-02), p. 441-451
    Details
    In: Infection and Immunity, American Society for Microbiology, Vol. 81, No. 2 ( 2013-02), p. 441-451
    Abstract: Blood-stage malaria vaccines that target single Plasmodium falciparum antigens involved in erythrocyte invasion have not induced optimal protection in field trials. Blood-stage malaria vaccine development has faced two major hurdles, antigenic polymorphisms and molecular redundancy, which have led to an inability to demonstrate potent, strain-transcending, invasion-inhibitory antibodies. Vaccines that target multiple invasion-related parasite proteins may inhibit erythrocyte invasion more efficiently. Our approach is to develop a receptor-blocking blood-stage vaccine against P. falciparum that targets the erythrocyte binding domains of multiple parasite adhesins, blocking their interaction with their receptors and thus inhibiting erythrocyte invasion. However, with numerous invasion ligands, the challenge is to identify combinations that elicit potent strain-transcending invasion inhibition. We evaluated the invasion-inhibitory activities of 20 different triple combinations of antibodies mixed in vitro against a diverse set of six key merozoite ligands, including the novel ligands P. falciparum apical asparagine-rich protein (PfAARP), EBA-175 (PfF2), P. falciparum reticulocyte binding-like homologous protein 1 (PfRH1), PfRH2, PfRH4, and Plasmodium thrombospondin apical merozoite protein (PTRAMP), which are localized in different apical organelles and are translocated to the merozoite surface at different time points during invasion. They bind erythrocytes with different specificities and are thus involved in distinct invasion pathways. The antibody combination of EBA-175 (PfF2), PfRH2, and PfAARP produced the most efficacious strain-transcending inhibition of erythrocyte invasion against diverse P. falciparum clones. This potent antigen combination was selected for coimmunization as a mixture that induced balanced antibody responses against each antigen and inhibited erythrocyte invasion efficiently. We have thus demonstrated a novel two-step screening approach to identify a potent antigen combination that elicits strong strain-transcending invasion inhibition, supporting its development as a receptor-blocking malaria vaccine.
    Type of Medium: Online Resource
    ISSN: 0019-9567 , 1098-5522
    URL: Article
    DOI: 10.1128/IAI.01107-12
    RVK:
    XA 10000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2013
    detail.hit.zdb_id: 1483247-1
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  • 4
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    Bacterially Expressed Full-Length Recombinant Plasmodium falciparum RH5 Protein Binds Erythrocytes and Elicits Potent Strain-Transcending Parasite-Neutralizing Antibodies
    American Society for Microbiology ; 2014
    In:  Infection and Immunity Vol. 82, No. 1 ( 2014-01), p. 152-164
    Details
    In: Infection and Immunity, American Society for Microbiology, Vol. 82, No. 1 ( 2014-01), p. 152-164
    Abstract: Plasmodium falciparum reticulocyte binding-like homologous protein 5 (PfRH5) is an essential merozoite ligand that binds with its erythrocyte receptor, basigin. PfRH5 is an attractive malaria vaccine candidate, as it is expressed by a wide number of P. falciparum strains, cannot be genetically disrupted, and exhibits limited sequence polymorphisms. Viral vector-induced PfRH5 antibodies potently inhibited erythrocyte invasion. However, it has been a challenge to generate full-length recombinant PfRH5 in a bacterial-cell-based expression system. In this study, we have produced full-length recombinant PfRH5 in Escherichia coli that exhibits specific erythrocyte binding similar to that of the native PfRH5 parasite protein and also, importantly, elicits potent invasion-inhibitory antibodies against a number of P. falciparum strains. Antibasigin antibodies blocked the erythrocyte binding of both native and recombinant PfRH5, further confirming that they bind with basigin. We have thus successfully produced full-length PfRH5 as a functionally active erythrocyte binding recombinant protein with a conformational integrity that mimics that of the native parasite protein and elicits potent strain-transcending parasite-neutralizing antibodies. P. falciparum has the capability to develop immune escape mechanisms, and thus, blood-stage malaria vaccines that target multiple antigens or pathways may prove to be highly efficacious. In this regard, antibody combinations targeting PfRH5 and other key merozoite antigens produced potent additive inhibition against multiple worldwide P. falciparum strains. PfRH5 was immunogenic when immunized with other antigens, eliciting potent invasion-inhibitory antibody responses with no immune interference. Our results strongly support the development of PfRH5 as a component of a combination blood-stage malaria vaccine.
    Type of Medium: Online Resource
    ISSN: 0019-9567 , 1098-5522
    URL: Article
    DOI: 10.1128/IAI.00970-13
    RVK:
    XA 10000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2014
    detail.hit.zdb_id: 1483247-1
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  • 5
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    Implications of Limits of Detection of Various Methods for Bacillus anthracis in Computing Risks to Human Health
    American Society for Microbiology ; 2009
    In:  Applied and Environmental Microbiology Vol. 75, No. 19 ( 2009-10), p. 6331-6339
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 75, No. 19 ( 2009-10), p. 6331-6339
    Abstract: Used for decades for biological warfare, Bacillus anthracis (category A agent) has proven to be highly stable and lethal. Quantitative risk assessment modeling requires descriptive statistics of the limit of detection to assist in defining the exposure. Furthermore, the sensitivities of various detection methods in environmental matrices are vital information for first responders. A literature review of peer-reviewed journal articles related to methods for detection of B. anthracis was undertaken. Articles focused on the development or evaluation of various detection approaches, such as PCR, real-time PCR, immunoassay, etc. Real-time PCR and PCR were the most sensitive methods for the detection of B. anthracis , with median instrument limits of detection of 430 and 440 cells/ml, respectively. There were very few peer-reviewed articles on the detection methods for B. anthracis in the environment. The most sensitive limits of detection for the environmental samples were 0.1 CFU/g for soil using PCR-enzyme-linked immunosorbent assay (ELISA), 17 CFU/liter for air using an ELISA-biochip system, 1 CFU/liter for water using cultivation, and 1 CFU/cm 2 for stainless steel fomites using cultivation. An exponential dose-response model for the inhalation of B. anthracis estimates of risk at concentrations equal to the environmental limit of detection determined the probability of death if untreated to be as high as 0.520. Though more data on the environmental limit of detection would improve the assumptions made for the risk assessment, this study's quantification of the risk posed by current limitations in the knowledge of detection methods should be considered when employing those methods in environmental monitoring and cleanup strategies.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.00288-09
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2009
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 6
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    Unmasking of CgYor1-Dependent Azole Resistance Mediated by Target of Rapamycin (TOR) and Calcineurin Signaling in Candida glabrata
    American Society for Microbiology ; 2022
    In:  mBio Vol. 13, No. 1 ( 2022-02-22)
    Details
    In: mBio, American Society for Microbiology, Vol. 13, No. 1 ( 2022-02-22)
    Abstract: In this study, 18 predicted membrane-localized ABC transporters of Candida glabrata were deleted individually to create a minilibrary of knockouts (KO). The transporter KOs were analyzed for their susceptibility toward antimycotic drugs. Although Cg YOR1 has previously been reported to be upregulated in various azole-resistant clinical isolates of C. glabrata , deletion of this gene did not change the susceptibility to any of the tested azoles. Additionally, Cg yor1 Δ showed no change in susceptibility toward oligomycin, which is otherwise a well-known substrate of Yor1 in other yeasts. The role of CgYor1 in azole susceptibility only became evident when the major transporter Cg CDR1 gene was deleted. However, under nitrogen-depleted conditions, Cg yor1 Δ demonstrated an azole-susceptible phenotype, independent of CgCdr1. Notably, Cg yor1Δ cells also showed increased susceptibility to target of rapamycin (TOR) and calcineurin inhibitors. Moreover, increased phytoceramide levels in Cg yor1 Δ and the deletions of regulators downstream of TOR and the calcineurin signaling cascade (Cg ypk1 Δ, Cg ypk2Δ , Cg ckb1 Δ, and Cg ckb2 Δ) in the Cg yor1 Δ background and their associated fluconazole (FLC) susceptibility phenotypes confirmed their involvement. Collectively, our findings show that TOR and calcineurin signaling govern CgYor1-mediated azole susceptibility in C. glabrata . IMPORTANCE The increasing incidence of Candida glabrata infections in the last 40 years is a serious concern worldwide. These infections are usually associated with intrinsic azole resistance and increasing echinocandin resistance. Efflux pumps, especially ABC transporter upregulation, are one of the prominent mechanisms of azole resistance; however, only a few of them are characterized. In this study, we analyzed the mechanisms of azole resistance due to a multidrug resistance-associated protein (MRP) subfamily ABC transporter, CgYor1. We demonstrate for the first time that CgYor1 does not transport oligomycin but is involved in azole resistance. Under normal growing conditions its function is masked by major transporter CgCdr1; however, under nitrogen-depleted conditions, it displays its azole resistance function independently. Moreover, we propose that the azole susceptibility due to removal of CgYor1 is not due to its transport function but involves modulation of TOR and calcineurin cascades.
    Type of Medium: Online Resource
    ISSN: 2150-7511
    URL: Article
    DOI: 10.1128/mbio.03545-21
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 2557172-2
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  • 7
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    Evaluation of Sample Recovery Efficiency for Bacteriophage P22 on Fomites
    American Society for Microbiology ; 2012
    In:  Applied and Environmental Microbiology Vol. 78, No. 22 ( 2012-11-15), p. 7915-7922
    Details
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 78, No. 22 ( 2012-11-15), p. 7915-7922
    Abstract: Fomites are known to play a role in the transmission of pathogens. Quantitative analysis of the parameters that affect sample recovery efficiency (SRE) at the limit of detection of viruses on fomites will aid in improving quantitative microbial risk assessment (QMRA) and infection control. The variability in SRE as a function of fomite type, fomite surface area, sampling time, application media, relative humidity (rH), and wetting agent was evaluated. To quantify the SRE, bacteriophage P22 was applied onto fomites at average surface densities of 0.4 ± 0.2 and 4 ± 2 PFU/cm 2 . Surface areas of 100 and 1,000 cm 2 of nonporous fomites found in indoor environments (acrylic, galvanized steel, and laminate) were evaluated with premoistened antistatic wipes. The parameters with the most effects on the SRE were sampling time, fomite surface area, wetting agent, and rH. At time zero (the initial application of bacteriophage P22), the SRE for the 1,000-cm 2 fomite surface area was, on average, 40% lower than that for the 100-cm 2 fomite surface area. For both fomite surface areas, the application medium Trypticase soy broth (TSB) and/or the laminate fomite predominantly resulted in a higher SRE. After the applied samples dried on the fomites (20 min), the average SRE was less than 3%. A TSB wetting agent applied on the fomite improved the SRE for all samples at 20 min. In addition, an rH greater than 28% generally resulted in a higher SRE than an rH less than 28%. The parameters impacting SRE at the limit of detection have the potential to enhance sampling strategies and data collection for QMRA models.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    URL: Article
    DOI: 10.1128/AEM.01370-12
    RVK:
    WA 15000
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2012
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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