title: Characterization of a serodiagnostic complement fixation antigen of Coccidioides posadasii expressed in the nonpathogenic Fungus Uncinocarpus reesii.
abstract: Coccidioides spp. (immitis and posadasii) are the causative agents of human coccidioidomycosis. In this study, we developed a novel system to overexpress coccidioidal proteins in a nonpathogenic fungus, Uncinocarpus reesii, which is closely related to Coccidioides. A promoter derived from the heat shock protein gene (HSP60) of Coccidioides posadasii was used to control the transcription of the inserted gene in the constructed coccidioidal protein expression vector (pCE). The chitinase gene (CTS1) of C. posadasii, which encodes the complement fixation antigen, was expressed using this system. The recombinant Cts1 protein (rCts1(Ur)) was induced in pCE-CTS1-transformed U. reesii by elevating the cultivation temperature. The isolated rCts1(Ur) showed chitinolytic activity that was identical to that of the native protein and had serodiagnostic efficacy comparable to those of the commercially available antigens in immunodiffusion-complement fixation tests. Using the purified rCts1(Ur), 74 out of the 77 coccidioidomycosis patients examined (96.1%) were positively identified by enzyme-linked immunosorbent assay. The rCts1(Ur) protein showed higher chitinolytic activity and slightly greater seroreactivity than the bacterially expressed recombinant Cts1. These data suggest that this novel expression system is a useful tool to produce coccidioidal antigens for use as diagnostic antigens.
authors.affiliation: Department of Medical Microbiology and Immunology, Medical College of Ohio, Toledo, Ohio, USA.
authors.forename: J-J
authors.initials: JJ
authors.lastname: Yu
authors.forename: T N
authors.initials: TN
authors.lastname: Kirkland
authors.forename: L K
authors.initials: LK
authors.lastname: Hall
authors.forename: J
authors.initials: J
authors.lastname: Wopschall
authors.forename: R C
authors.initials: RC
authors.lastname: Smith
authors.forename: C-Y
authors.initials: CY
authors.lastname: Hung
authors.forename: X
authors.initials: X
authors.lastname: Chen
authors.forename: E
authors.initials: E
authors.lastname: Tarcha
authors.forename: P W
authors.initials: PW
authors.lastname: Thomas
authors.forename: G T
authors.initials: GT
authors.lastname: Cole
mesh_terms: D000946:Antigens, Fungal / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D002686:Chitin / Q000378:metabolism; D003045:Coccidioides / Q000276:immunology*; D003047:Coccidioidomycosis / Q000175:diagnosis; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005658:Fungi / Q000378:metabolism*; D015870:Gene Expression; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012698:Serologic Tests
chemical_list: D000946:Antigens, Fungal; D011994:Recombinant Proteins; D002686:Chitin