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Demosthenes Pappagianis - One of the best experts on this subject based on the ideXlab platform.

  • coccidioidomycosis Complement Fixation titer trends in the age of antifungals
    Journal of Clinical Microbiology, 2018
    Co-Authors: Ian Howard Mchardy, Demosthenes Pappagianis, Bao Tran N Dinh, Sarah Waldman, Ethan R Stewart, Derek J Bays, George Richard Thompson
    Abstract:

    Coccidioidomycosis is associated with a broad spectrum of illness severity, ranging from asymptomatic or self-limited pulmonary infection to life-threatening manifestations of disseminated disease. Serologic studies before the widespread availability of antifungals established current understanding of serologic kinetics and dynamics. Chart histories and Complement Fixation (CF) titer trends were analyzed for 434 antifungal-treated coccidioidomycosis patients, who were classified by three infectious disease physicians as having either pulmonary uncomplicated coccidioidomycosis (PUC) (n = 248), pulmonary chronic coccidioidomycosis (PCC) (n = 64), disseminated coccidioidomycosis (DC) not including meningitis (n = 86), or coccidioidal meningitis (CM) (n = 36). The median maximal CF titers were 1:4 for PUC patients, 1:24 for PCC patients, 1:128 for DC patients, and 1:32 for CM patients. Approximately 25.4% of PUC patients, 6.2% of PCC patients, 2.3% of DC patients, and 8.3% of CM patients did not develop detectable titers during the study period. Maximal titers developed a mean of 31 days (95% confidence interval [CI], 13 to 50 days) after initial serologic positivity, with no significant differences between groups. Serologic recurrence occurred in 9% of PUC patients, 36% of PCC patients, 50% of DC patients, and 52% of CM patients. Median titer improvement rates were 91 days/dilution for PUC patients, 112 days/dilution for PCC patients, 136 days/dilution for DC patients, and 146 days/dilution for CM patients. Receiver operating characteristic (ROC) analysis revealed that CF testing retains moderate classification value for disseminated infections (area under the curve [AUC], 0.82 [95% CI, 0.78 to 0.87]) and complicated infections (AUC, 0.82 [95% CI, 0.77 to 0.86]). A suitable cutoff value for complicated infections is ≥1:32. Findings update serologic parameters that are relevant for clinical assessment of coccidioidomycosis patients in the triazole era.

  • cloning and expression of the Complement Fixation antigen chitinase of coccidioides immitis
    Infection and Immunity, 1996
    Co-Authors: C R Zimmermann, Suzanne M. Johnson, Gregory W Martens, A G White, Demosthenes Pappagianis
    Abstract:

    A chitinase had been isolated from the culture filtrates of Coccidioides immitis endosporulating spherules and from hyphae and shown to be the coccidioidal Complement Fixation (CF) and immunodiffusion-CF antigen. In the present study, we made use of our previously determined amino-terminal (N-terminal) sequence of the CF-chitinase to design degenerate oligonucleotide primers and to amplify and sequence a PCR product that coded for the N-terminal portion of the CF-chitinase. The PCR product was used as a hybridization probe to screen a developing spherule-(lambda)ZAP cDNA library, and three hybridizing clones were selected. These clones were converted into their pBluescript expression plasmid form in Escherichia coli and induced to express their recombinant proteins. Lysate from only one clone, pCTS 4-2A, yielded an enzymatically functional CF-chitinase and a line of identity with control immunodiffusion-CF-positive antigen. The pCTS 4-2A insert was sequenced and found to contain a deduced open reading frame coding for a 427-amino-acid polypeptide with an approximate molecular weight of 47 kDa. When purified by a chitin adsorption-desorption method, the recombinant protein exhibited virtually identical characteristics to those of the original C. immitis CF-chitinase. Nondenaturing gels of the pCTS 4-2A E. coli lysates and the purified C. immitis and recombinant CF-chitinase revealed proteins that had chitinase activity and similar relative electrophoretic mobilities. The appearance and relative levels of hybridizing RNA from the developing spherules-endospores (SEs) and hyphae correlated with the appearance or presence and level of CF-chitinase enzyme activity found in SEs culture filtrate and in cellular extracts of developing SE and hyphae. Thus, a functional recombinant CF-chitinase antigen was produced in E. coli and was used in serological diagnostic applications. These results also suggest a functional role for this chitinase in SE development and maturation.

  • comparative evaluation of commercial premier eia and microimmunodiffusion and Complement Fixation tests for coccidioides immitis antibodies
    Journal of Clinical Microbiology, 1995
    Co-Authors: Leo Kaufman, A S Sekhon, N Moledina, M Jalbert, Demosthenes Pappagianis
    Abstract:

    A total of 409 serum and cerebrospinal fluid specimens from human subjects with proven coccidioidomycosis, with other infections, or with no apparent illness were tested for antibodies to Coccidioides immitis by the Premier EIA (Meridian Diagnostics, Inc., Cincinnati, Ohio), which tests for immunoglobulin G (IgG) and IgM responses to coccidioidal antigens, and by the conventional Complement Fixation (CF) or immunodiffusion (ID) assays for antibodies corresponding to those detected by the tube precipitin (TP) or CF tests. Of the 409 specimens, 47 were from persons with confirmed coccidioidomycosis and all were positive for C. immitis antibodies in IDCF tests and enzyme immunoassays (EIAs) for both IgG and IgM. The EIA for detecting both IgG and IgM antibodies proved to be sensitive for detecting coccidioidomycosis case sera positive by the IDCF, IDTP, and CF tests. Maximal sensitivity for diagnosing coccidioidomycosis is dependent upon detection of both IgG and IgM antibodies in the EIA. The EIA, however, was not absolutely specific, since some sera from patients with confirmed blastomycosis and some from patients with noncoccidioidal disease produced false-positive reactions.

  • amino terminal sequence analysis of the coccidioides immitis chitinase immunodiffusion Complement Fixation protein
    Infection and Immunity, 1993
    Co-Authors: Suzanne M. Johnson, C R Zimmermann, Demosthenes Pappagianis
    Abstract:

    A chitinase isolated from Coccidioides immitis was subjected to amino-terminal protein sequence analysis. The resulting 18-amino-acid sequence was compared with the previously reported amino acid sequence of coccidioidal immunodiffusion-Complement Fixation (IDCF) antigen. From the homology of the two sequences, the results support the identification of the IDCF antigen with a chitinase.

  • the coccidioidal Complement Fixation and immunodiffusion Complement Fixation antigen is a chitinase
    Infection and Immunity, 1992
    Co-Authors: Suzanne M. Johnson, Demosthenes Pappagianis
    Abstract:

    Culture filtrates and autolysates of Coccidioides immitis have provided suitable crude antigens for the serodiagnosis and prognosis of coccidioidomycosis. One of these, a heat-labile antigen which participates in the immunodiffusion reaction corresponding to the Complement Fixation reaction (IDCF), has been characterized as a 110-kDa native protein that, when subjected to reducing conditions and heat, yields a 48-kDa component. The present report provides serologic and biochemical evidence that this antigen is a chitinase. This chitinase, isolated from 48-h culture filtrate of the spherule-endospore-phase C. immitis by affinity adsorption to chitin, formed a line of identity with the IDCF reference antigen and participated in the Complement Fixation reaction with human serum. It lost its enzymatic as well as antigenic activity when heated, but when not heated it retained its enzymatic activity even when precipitated with coccidiodal antibody present in human serum. This chitinase represents a significant serodiagnostic substance and may be important in the morphogenesis of C. immitis. Images

Suzanne M. Johnson - One of the best experts on this subject based on the ideXlab platform.

  • cloning and expression of the Complement Fixation antigen chitinase of coccidioides immitis
    Infection and Immunity, 1996
    Co-Authors: C R Zimmermann, Suzanne M. Johnson, Gregory W Martens, A G White, Demosthenes Pappagianis
    Abstract:

    A chitinase had been isolated from the culture filtrates of Coccidioides immitis endosporulating spherules and from hyphae and shown to be the coccidioidal Complement Fixation (CF) and immunodiffusion-CF antigen. In the present study, we made use of our previously determined amino-terminal (N-terminal) sequence of the CF-chitinase to design degenerate oligonucleotide primers and to amplify and sequence a PCR product that coded for the N-terminal portion of the CF-chitinase. The PCR product was used as a hybridization probe to screen a developing spherule-(lambda)ZAP cDNA library, and three hybridizing clones were selected. These clones were converted into their pBluescript expression plasmid form in Escherichia coli and induced to express their recombinant proteins. Lysate from only one clone, pCTS 4-2A, yielded an enzymatically functional CF-chitinase and a line of identity with control immunodiffusion-CF-positive antigen. The pCTS 4-2A insert was sequenced and found to contain a deduced open reading frame coding for a 427-amino-acid polypeptide with an approximate molecular weight of 47 kDa. When purified by a chitin adsorption-desorption method, the recombinant protein exhibited virtually identical characteristics to those of the original C. immitis CF-chitinase. Nondenaturing gels of the pCTS 4-2A E. coli lysates and the purified C. immitis and recombinant CF-chitinase revealed proteins that had chitinase activity and similar relative electrophoretic mobilities. The appearance and relative levels of hybridizing RNA from the developing spherules-endospores (SEs) and hyphae correlated with the appearance or presence and level of CF-chitinase enzyme activity found in SEs culture filtrate and in cellular extracts of developing SE and hyphae. Thus, a functional recombinant CF-chitinase antigen was produced in E. coli and was used in serological diagnostic applications. These results also suggest a functional role for this chitinase in SE development and maturation.

  • amino terminal sequence analysis of the coccidioides immitis chitinase immunodiffusion Complement Fixation protein
    Infection and Immunity, 1993
    Co-Authors: Suzanne M. Johnson, C R Zimmermann, Demosthenes Pappagianis
    Abstract:

    A chitinase isolated from Coccidioides immitis was subjected to amino-terminal protein sequence analysis. The resulting 18-amino-acid sequence was compared with the previously reported amino acid sequence of coccidioidal immunodiffusion-Complement Fixation (IDCF) antigen. From the homology of the two sequences, the results support the identification of the IDCF antigen with a chitinase.

  • the coccidioidal Complement Fixation and immunodiffusion Complement Fixation antigen is a chitinase
    Infection and Immunity, 1992
    Co-Authors: Suzanne M. Johnson, Demosthenes Pappagianis
    Abstract:

    Culture filtrates and autolysates of Coccidioides immitis have provided suitable crude antigens for the serodiagnosis and prognosis of coccidioidomycosis. One of these, a heat-labile antigen which participates in the immunodiffusion reaction corresponding to the Complement Fixation reaction (IDCF), has been characterized as a 110-kDa native protein that, when subjected to reducing conditions and heat, yields a 48-kDa component. The present report provides serologic and biochemical evidence that this antigen is a chitinase. This chitinase, isolated from 48-h culture filtrate of the spherule-endospore-phase C. immitis by affinity adsorption to chitin, formed a line of identity with the IDCF reference antigen and participated in the Complement Fixation reaction with human serum. It lost its enzymatic as well as antigenic activity when heated, but when not heated it retained its enzymatic activity even when precipitated with coccidiodal antibody present in human serum. This chitinase represents a significant serodiagnostic substance and may be important in the morphogenesis of C. immitis. Images

  • the coccidioidal Complement Fixation and immunodiffusion Complement Fixation antigen is a chitinase
    Infection and Immunity, 1992
    Co-Authors: Suzanne M. Johnson, Demosthenes Pappagianis
    Abstract:

    Culture filtrates and autolysates of Coccidioides immitis have provided suitable crude antigens for the serodiagnosis and prognosis of coccidioidomycosis. One of these, a heat-labile antigen which participates in the immunodiffusion reaction corresponding to the Complement Fixation reaction (IDCF), has been characterized as a 110-kDa native protein that, when subjected to reducing conditions and heat, yields a 48-kDa component. The present report provides serologic and biochemical evidence that this antigen is a chitinase. This chitinase, isolated from 48-h culture filtrate of the spherule-endospore-phase C. immitis by affinity adsorption to chitin, formed a line of identity with the IDCF reference antigen and participated in the Complement Fixation reaction with human serum. It lost its enzymatic as well as antigenic activity when heated, but when not heated it retained its enzymatic activity even when precipitated with coccidiodal antibody present in human serum. This chitinase represents a significant serodiagnostic substance and may be important in the morphogenesis of C. immitis.

Amy L Leber - One of the best experts on this subject based on the ideXlab platform.

Rosanna Adone - One of the best experts on this subject based on the ideXlab platform.

  • development of a sterne based Complement Fixation test to monitor the humoral response induced by anthrax vaccines
    Frontiers in Microbiology, 2016
    Co-Authors: Rosanna Adone, Massimiliano Francia, Michela Sali, Michela Iatarola, Adelia Donatiello, Antonio Fasanella
    Abstract:

    Anthrax is a zoonotic disease caused by Bacillus anthracis spore-forming bacterium. Since it is primarily a disease of animals, the control in animals and humans depends on the prevention in livestock, principally cattle, sheep and goats. Most veterinary vaccines utilize the toxigenic, uncapsulated (pXO1+/pXO2–) B.anthracis strain 34F2 which affords protection through the production of neutralizing antibodies directed to the toxin components Protective Antigen (PA), Lethal Factor (LF) and Edema Factor (EF). The titration of specific antibodies in sera of vaccinated animals is crucial to evaluate the efficacy of the vaccination and to obtain epidemiological information for an effective anthrax surveillance. In this study, we developed a Sterne-based Complement Fixation Test to detect specific antibodies induced in animals vaccinated with Sterne 34F2 . We assessed its efficacy in laboratory animals and under field conditions by monitoring the humoral response induced by vaccination in cattle. The results indicated that the Sterne-based CFT is able to identify vaccinated animals with a good sensitivity and specificity offering many benefits especially with regard to costs, standardization and reproducibility of the assay procedure.

  • Brucella melitensis B115-based Complement Fixation test to detect antibodies induced by Brucella rough strains.
    Journal of Applied Microbiology, 2008
    Co-Authors: Rosanna Adone, Massimiliano Francia, Franco Ciuchini
    Abstract:

    Aims:  To assess the efficiency of a Brucella melitensis B115 rough strain, naturally devoid of antiComplementary activity, used as antigen in a Complement Fixation test (CFT) to detect antibodies induced by Brucella strains with rough phenotype, such as Brucella abortus RB51, Brucella ovis and Brucella canis. Methods and Results:  Complement Fixation testing was performed on sera from RB51-vaccinated cattle and buffaloes, B. ovis-infected sheep and B. canis-infected dogs using B115, RB51 and the hot saline extract (HSE) as antigens. The B115-based CFT proved highly sensitive and specific in detecting rough antibodies and its efficiency was comparable with that of RB51 and HSE-based CFT. Conclusions: Brucella melitensis B115 can be successfully used as an antigen in CFT to detect antibodies induced by Brucella rough strains. Significance and Impact of the Study: Brucella melitensis B115 antigen may represent an improvement over Brucella rough strains for Brucella antibody detection by CFT, thus enhancing the efficiency of brucellosis surveillance systems. Owing to the absence of antiComplementary activity, it does not require particular growth conditions or modifications and can be accurately standardized. The B115-based CFT may constitute a suitable supplementary test for the diagnosis of human infections owing to rough Brucellae.

  • Complement Fixation test to assess humoral immunity in cattle and sheep vaccinated with brucella abortus rb51
    Clinical and Vaccine Immunology, 1999
    Co-Authors: Rosanna Adone, Franco Ciuchini
    Abstract:

    The live attenuated Brucella abortus strain RB51 is a rifampin-resistant, lipopolysaccharide (LPS) O-chain-deficient mutant of virulent B. abortus 2308. The reduced O-chain content in RB51 prevents this bacterium from inducing antibodies detectable by the conventional serologic tests for bovine brucellosis diagnosis that mainly identify antibodies to LPS. The absence of available serologic tests for RB51 also complicates the diagnosis of possible RB51 infections in humans exposed to this strain. The purpose of this study was to evaluate the suitability of a Complement Fixation (CF) test performed with the rough strain B. abortus RB51, previously deprived of antiComplementary activity, in detecting anti-B. abortus RB51 antibodies in cattle and sheep experimentally vaccinated with this strain. The results of this study showed that a CF test with RB51 as the antigen is able to specifically detect antibodies following RB51 vaccination in cattle and sheep. In addition, this method could be a useful tool for detecting B. abortus RB51 infection in humans.

Franco Ciuchini - One of the best experts on this subject based on the ideXlab platform.

  • Brucella melitensis B115-based Complement Fixation test to detect antibodies induced by Brucella rough strains.
    Journal of Applied Microbiology, 2008
    Co-Authors: Rosanna Adone, Massimiliano Francia, Franco Ciuchini
    Abstract:

    Aims:  To assess the efficiency of a Brucella melitensis B115 rough strain, naturally devoid of antiComplementary activity, used as antigen in a Complement Fixation test (CFT) to detect antibodies induced by Brucella strains with rough phenotype, such as Brucella abortus RB51, Brucella ovis and Brucella canis. Methods and Results:  Complement Fixation testing was performed on sera from RB51-vaccinated cattle and buffaloes, B. ovis-infected sheep and B. canis-infected dogs using B115, RB51 and the hot saline extract (HSE) as antigens. The B115-based CFT proved highly sensitive and specific in detecting rough antibodies and its efficiency was comparable with that of RB51 and HSE-based CFT. Conclusions: Brucella melitensis B115 can be successfully used as an antigen in CFT to detect antibodies induced by Brucella rough strains. Significance and Impact of the Study: Brucella melitensis B115 antigen may represent an improvement over Brucella rough strains for Brucella antibody detection by CFT, thus enhancing the efficiency of brucellosis surveillance systems. Owing to the absence of antiComplementary activity, it does not require particular growth conditions or modifications and can be accurately standardized. The B115-based CFT may constitute a suitable supplementary test for the diagnosis of human infections owing to rough Brucellae.

  • Complement Fixation test to assess humoral immunity in cattle and sheep vaccinated with brucella abortus rb51
    Clinical and Vaccine Immunology, 1999
    Co-Authors: Rosanna Adone, Franco Ciuchini
    Abstract:

    The live attenuated Brucella abortus strain RB51 is a rifampin-resistant, lipopolysaccharide (LPS) O-chain-deficient mutant of virulent B. abortus 2308. The reduced O-chain content in RB51 prevents this bacterium from inducing antibodies detectable by the conventional serologic tests for bovine brucellosis diagnosis that mainly identify antibodies to LPS. The absence of available serologic tests for RB51 also complicates the diagnosis of possible RB51 infections in humans exposed to this strain. The purpose of this study was to evaluate the suitability of a Complement Fixation (CF) test performed with the rough strain B. abortus RB51, previously deprived of antiComplementary activity, in detecting anti-B. abortus RB51 antibodies in cattle and sheep experimentally vaccinated with this strain. The results of this study showed that a CF test with RB51 as the antigen is able to specifically detect antibodies following RB51 vaccination in cattle and sheep. In addition, this method could be a useful tool for detecting B. abortus RB51 infection in humans.